ISSN:0144-557X    

DOI:10.1039/AP98017FX044

出版商:RSC    

年代:1980

数据来源: RSC

ISSN:0144-557X    

DOI:10.1039/AP98017BX046

出版商:RSC    

年代:1980

数据来源: RSC

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ANPRDI 17(12) 507-560 (1980) ISSN 01 44-557X D ecem ber 1980 Analvtical Proceedinus w - Proceedings of the Analytical Division of The Royal Society of Chemistry AD President L. S . Bark Hon. Secretary P. G. W. Cobb Hon. Assistant Secretary D. I. Coomber, O.B.E. Hon. Publicity and Public Relations Officer Dr. A. Townshend, Department of Chemistry, University of Hull, Hull, HU6 7RX Secretary Miss P. E. Hutchinson Editor, Analyst and Analytical Proceedings P. C. Weston Assistant Editors Mrs. J. Brew, R. W. Hazell, R. A. Young Publication of Analytical Proceedings is the responsi- bility of the Analyst Publications Committee: J. M. Ottaway (Chairman) W. H. C. Shaw H. J. Cluley D. Simpson ‘P. Gray J. M. Skinner J. N. Miller A. Townshend G. E. Penketh ‘P. C. Weston T. B. Pierce J.Whitehead ‘Ex officio members All editorial matter should be addressed to: The Editor, Analytical Proceedings, The Royal Society of Chemistry, Burlington House, Piccadilly, London, W1 V OBN. Telephone 01 -734 9864. Telex 268001. Advertisements: Advertising Department, The Royal Society of Chemistry, Burlington House, Piccadilly, London, W1 V OBN. Telephone 01 -734 9864. 0 The Royal Society of Chemistry 1980 Editorial Separate Subscription to Analvtical Proceedings In the course of the last year the RSC, through its Analyst Publications Committee, has taken a number of steps to increase the appeal of Analytical Proceedings. In addition to such changes as the introduction of a coloured cover, the acceptance of advertisements from outside sources (advertisements have always been en- couraged from the various subject groups and regions of the Analytical Division), the inclusion of a Reader Enquiry Service form and the intro- duction of “mini” reviews of some of the publications received for possible review in The Analyst, an attempt has been made to improve the service offered to members of the society by posting members’ copies separately in advance of The Analyst and Analytical Abstvacts.Al- though printing difficulties in the middle of the year thwarted this attempt for a while, members should now be noticing the effect of this separate posting. In January 1981 the Society will make a change which is intended to improve the service offered to non-member subscribers. As a new venture, Analytical Proceedings will be available to non-members separately from the other analytical journals at a cost of i30.00 in the UK and Eire, $70.50 in the USA and i31.50 in the Rest of the World.A “package” of all three analytical journals at an attractive price will still be offered to institutions and other sub- scribers who need to take all three, and it will still be possible to obtain Analytical A bstracts separately. It is hoped that by offering Analytical Proceedings separately the Society will enable current individual non-member‘ subscribers to continue to take the journal and will attract new readers. Analytical Proceedings increased sub- stantially in size in 1980 and the scope of the contents was also broadened considerably so that the Society feels confident that the 1981 prices represent good value and will be attractive to individual subscribers in all disciplines of analytical chemistry. Subscription details appear on p. 554 of this issue. R. A. YOUNG 507508 ADVANCES I N APPLIED HPLC Anal. Proc. Obituary: Dr. J m Km Foreman Il7e deeply regret to announce the death on November 14th of Dr. J . K. Foreman, Honorary Treasurer of the Analytical Division.

ISSN:0144-557X    

DOI:10.1039/AP9801700507

出版商:RSC    

年代:1980

数据来源: RSC

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508 ADVANCES I N APPLIED HPLC Anal. Proc. Advances in Applied HPLC The following are summaries of six of the papers presented at a Meeting of the Scottish Region, the Chromatography and Electrophoresis and Joint Pharmaceutical Analysis Groups and the Association of Clinical Biochemists held on February ZOth, 1980, at the University of Edinburgh. Optimum Conditions for High-performance Liquid Chromatography on the Preparative Scale A. W. J. de Jong, J. C. Smit, H. Poppe and J. C. Kraak Laboratory.for Aizalytical Chemistry, University of Amsterdam, Nieuwe Achfergracht 166, 1018 LV L' Aniskrdam, The Netherlands In preparative liquid chromatography the throughput (grams per second) is most often used as a performance parameter. The maximum obtainable throughput will be mainly deter- mined by (i) geometric and kinetic parameters (column dimensions, particle size, injection mode and eluting agent velocity), (ii) parameters of the selected phase system (selectivity factor and usable range of the distribution isotherm) and (iii) problem-specific parameters, such as composition of the sample and the specification with respect to yield and purity of the product obtained.One of the most important parameters for increasing the throughput is the selectivity factor, as was shown by Wehrli et aZ.l Optimisation of the throughput by examination of the selectivity has to be carried out first. However, when separating complex mixtures one is often confronted with the fact that the selectivity factor of one pair of compounds can be improved only at the expense of the selectivity factor of another pair.In such instances one needs high plate numbers in order to avoid contamination of the products by closely eluting compounds. The only way of improving the throughput is then obtained by adjust- ment of the geometric and kinetic factors, taking into account limitations such as the plate number required, available pressure drop and maximum feasible cross-section of the column. It can be shown2 that as soon as the phase system and column diameter have been chosen and when the particle size and column length are adapted to the required plate number and pressure limitation, the throughput is proportional to (./It)+, where v is the reduced velocity and h the reduced plate height.3 This result shows that one must work under conditions where v / h is as high as possible.This can be realised when one works on the ascending branch of the H / v curve, where the major part of the plate height is due to intra-particle mass transfer. This points to particle sizes of 20-50 pm and long columns, as is shown in Fig. 1. Another important effect that can influence the throughput significantly, especially when purity specifications are stringent, is peak tailing in the lower parts of the elution curves. This peak tailing is determined by irregularities of the packing near the walls of the column, radial temperature gradients and the method of sample introduction. Resolution and plate- height calculations from peak-width measurements at the higher parts of the elution curve (e.g., at 0.6 of the maximum peak height) ignore the tailing in the lower parts and can be misleading in throughput studies.A more realistic characterisation of the performance of preparative columns is obtained when these quantities are calculated from the peak width in the lower parts of the elution curves (e.g., at 0.1 of the maximum peak height). The peak tailing caused by the method of sample introduction at 0.1 of the peak height can be significantly improved by application of the so-called flow surrounding conical disperser column t ~ p , ~ , ~ shown schematically in Fig. 2. In particular, when separating complex mixtures, with small selectivity factors requiring high plate numbers, this type of injection mode is of great value.December, 1980 ADVANCES I N APPLIED HPLC 509 P = 80 bar N = 8000 P = 200 bar N = 8000 P = 60 bar 10 20 Y 30 Fig.1. Graph of (u/h)3 uemm u . Adsorbent: silica gel SI 60; 0, 5-8 pm ; a, 20-25 pm. Mobile phase : 2,2,4-trimethylpentane + 1 yo I.'/ L' butan-1-01. Solute : 2,4-dimethylphenol. Another important factor that affects the throughput is Pump 1 Injection valve Glass beads 25-40 lcm Fig. 2. Schematic representa- tion of the flow surrounding conical column top terminator. the shaDe of the distribution isotherm (i.e.,-the linear range). In practice one sh&dd inject amouLts of sample that are sufficiently large that two neighbouring peaks just start to overlap as a result of the extra peak broadening and peak distortion caused by non-linearity of the isotherm. How much sample can be injected depends on the linear range of the distribution isotherm and on the resolution required.It can be shown that it is the column outlet concentration and the deviation in linearity in the isotherm existing at that concentration that determine peak distortion and overload broadening. For the case of ideal chromatography (i.e., dispersionless, with an infinite plate number), theories already formulated in the 1940s and, for example, reviewed and reformulated by Huber and Gerrit~e,~ allow this conclusion. When the sample is injected as a spike,5 and even for a variety of other input conditions,6 the following equation holds : where cim = the concentration in the mobile phase of component i, cis = the concentration in the stationary phase of component i and 4 = the phase ratio.For linear isotherms the differential in equation (1) is the distribution constant Ki and equation (1) becomes the familiar expression ki = tRO (l + 4Ki) = t R 0 (l + K i ) .. .. * (2) where K$ = capacity factor of component i, yielding the same tR, for every cim value, that is, the spike input elutes as a spike at time tRi. For non-linear isotherms the equation gives different tRi values for various cim values. For the more usual case of a convex isotherm, theory predicts an elution curve with a sharp front, and a tail described by equation (1); for every concentration value cim the derivative of the isotherm at that concentration determines the tRi value (see Fig. 3). This means that the total amount of the compound i is spread out over a tR6 range, and the upper and lower limits of this tRi range are510 ADVANCES I N APPLIED HPLC Anal.Proc. i.e., the limits of this range are determined by the derivative of the isotherm for the lowest and highest values of the column outlet concentration. Note that the elution function is not at all influenced by the concentrations occurring at the top of the column; also, the shape of the isotherm at these much higher initial concentrations is irrelevant, and the phenomena occurring there are apparently “forgotten” in the following transport process. Now, for real, non-ideal, finite plate count situations we cannot solve the equation for the transport process, and therefore we cannot give an expression for the elution function. However, both experiment and computer simulation studies’ show that the outlet concentra- tion is also the relevant parameter.The experimental illustration is given in Fig. 4, where for two injection volumes for the same amount (but avoiding volume overload; Vjnj < uVi) identical elution functions result. Studies of optimisation of preparative liquid chromato- graphy2*s were therefore undertaken with the assumption that only outlet concentrations are important for the concentration overload. T E (5 tR Fig. 3. Influence of the amount of sample injected on the peak shape and the retention time a t the peak maximum. Column: 10 x 1 cm, filled with 5-pm silica (SI 60). Mobile phase : dichloromethane. Solute : phenol. Injection volume: 50 pl. fR Fig. 4. Influence of the volume in which the sample is injected on the peak shape Conditions as in Fig.3 except injection volume (50 or 600 pl). Injected anions of phenol: 400 pg. A further problem is which condition is preferable for preparative liquid chromatography : that of a highly efficient column with high concentration overload, leading to highly asym- metric peaks practically identical with those predicted by the ideal model, on the one hand, or a less efficient column, where peak broadening is determined predominantly by the dispersion in the column, on the other. Qualitatively, it can be derived easily that an intermediate situation is optimum with respect to throughput. If dispersion is still exclusively responsible for peak width (overlap), the throughput can be increased by increasing the concentration load.If, on the other hand, the peak width is determined exclusively by the concentration overload effect, the throughout can be increased by decreasing the plate number (e.g., larger particles in the same column would allow much higher flow-rates). This is illustrated in Fig. 5 , resulting from computer simulation studies.67’ The same amount was injected on columns with plate number varying from 250 to infinity (dispersionless). It can be seen that the peak width decreases only slightly when plate numbers are high (> 2000). Hence in this instance plate numbers in excess of about 2000 are essentially wasted, and detrimental to through- put, as the realisation of high plate numbers is very time c~nsuming.~ An effective peak width, b,.,,, was calculated from the curves in Fig.5 as the interval within which 95% of the total amount elutes. In Fig. 6 these bO1+, values are plotted against the plate number, N . For high plate numbers the peak width IS essentially constant. As a conclusion, we suggest the following strategy for solving a preparative liquid- chromatographic problem. 1. Find the phase system(s) that yield sufficient selectivity, reasonable retention andDecember, 1980 ADVANCES I N APPLIED HPLC 51 1 8.00 7.00 6.00 1 5.00 4 4.00 N X 3.00 2.00 6.00 6.50 7.00 7.50 8.00 8.50 9.00 9.50 10.00 t,/s x lo-’ __+ Elution functions obtained by simulation of the chromatographic transport with constant injected amount and varying plate number. high loadability for the compounds occurring in the mixture. As in analytical liquid chromatography, this is the most difficult step.Decide on the operating mode, i.e., decide between the following possibilities and/or corn binat ions : (i) single column, no recycling; (ii) single column, partial collection of compounds and recycling of mixture fractions (peak shaving and recycle) ; (iii) niulti-column operation. These decisions should be made on the basis of the requirements with respect to purity and yield, the composition of the sample and the performance of the phase systems. From (2), the acceptable overlap in critical parts of the chromatogram(s) can be derived. The classical resolution equation Fig. 5. 2. 3. can be used to calculate the required plate number corresponding to that overlap, for the case of infinite dilution. The actual specified plate number of the columns should then be 0 ’ I 5 000 N 10 000 Fig.6. Peak width, comprising 95% of the area, as a function of the plate number as obtained from the data in Fig. 5.Anal. Proc. increased somewhat in order to allow extra peak broadening due to overload, without violating the purity specification. Design a column(s) (particle size dimensions, velocity) with this specified plate number, allowing a flow-rate as high as possible. Load the system with increasing amounts of sample until the overlap surpasses the specification. In the resulting situation, peak broadening due to dispersion and that due to overload will be of about the same magnitude. 512 ADVANCES I N APPLIED HPLC 4. 1. 2. 3. 4. 5. 6. 7. 8. References W'ehrli, A., Hermann, U., and Huber, J .I;. K., J . Chvonzatogv., 1976, 125, 59. de Jong, A. W. J., Poppe, H., and Kraak, J. C., J . Chvoinalogv., 1958, 148, 147. Knox, J . H., and Salecm, M., .J. Chvomatogv. Sci., 1969, 7, 614. Webber, T. J . N., and McKerrell, E. H., .I. Chvomatogv., 1976, 122, 243. Huber, J. F. K., and Gerritse, K. G., J . Chromatogr., 1971, 58, 137. Smit, J . C., Smit, H. C., and de Jager, E. M., Anal. Chim. A c f a , Comput. Tech. Optim., 1980, 122, 1 . Smit, J. C., Smit, H. C., and de Jager, E. M., Anal. Chim. A c f a , Comput. Tech. Optirn., 1980, 122, 151. de Jong, A. W. J., Poppe, H., Kraak, J . C., and Nooitgcdagt, F., J . Chromatogr., 1980, 193, 181. Derivative and Multi-channel Detectors in High-performance Liquid Chromatography Anthony F. Fell Department of Pharniacy, Hcriot- Watt University, Edinbuvgh, E l i 1 2H J Although the technology and applications of high-performance liquid chromatography (HPLC) have developed at a faster rate than for any other analytical technique during the past decade, the detector still remains the factor that limits ultimate sensitivity and flexi- bility.Most devices based on the measurement of absorbed or emitted radiation have moderate sensitivity and are not universal in their response. The recent emergence of reliable rapid scanning spectrometers based on optical multi-channel detectors1 offers an alternative means for more flexible detection in HPLC, although this approach is still limited to analytes with appreciable absorption or fluorescence in the ultraviolet - visible spectrum.Data from optical multi-channel spectrometers may be manipulated by computer in various ways, either on- or off-line, to enhance resolution, specificity and sensitivity in HPLC. Among these methods, the presentation of three-dimensional A , A, t chromatograms and the first derivative (dA/dA) have been proposed.2 In this paper, these and other novel approaches to resolution enhancement in HPLC are described, including the technique where the second or fourth derivatives of the elution profile with respect to time (d2A/dt2, d4A/dt4) are genera- ted at a stationary ~avelength.~ The properties of this and other derivative functions are discussed with reference to their analytical potential when combined with multi-channel detectors in HPLC. Multi - channel Detectors Denton et u Z .~ gave an early demonstration of the greater flexibility offered by the rapid scanning detector in HPLC. Using a commercial oscillating galvanometer mirror with grating dispersion optics and a fast-response photomultiplier, three-dimensional plots of A , A, t chromatograms for model systems were obtained, while the normal elution chromatogram could be presented at stationary wavelengths optimised for each component. The use of the vidicon TV tube (Fig. 1) for detection in HPLC, as suggested by R ~ g e r s , ~ was later evaluated by McDowell and Pardue,"' who used a grating polychromator to disperse the eluate transmission spectrum across the front surface of a silicon-intensified vidicon (SIT) detector. In any one plane, several hundred photosensitive diode junctions act as spatially separated spectral resolution elements, each corresponding to a specific wavelength and monitored by a continuously scanning electron beam.l?* Incident radiation leads to the loss of charge in the diode, for which the re-charging current by electron beam is directly pro- portional to the incident radiation intensity at that wavelength.Hence, the entire spectrumDecember, 1980 ADVANCES I N APPLIED HPLC 513 is registered in the computer memory and can be manipulated in several ways. McDowell and Pardue explored the multi-wavelength capability of this detector using ensemble averaging to improve the signal to noise ratio (SNR), and successfully deconvoluted over- lapping chromatographic peaks in two pharmaceutical preparations (acetaminophen and phenylpropanolamine hydrochloride ; acetaminophen and caffeine) by the multi-wavelength matrix method.More recently, Klatt9 has developed criteria and computer algorithms for an SIT detector in HPLC that enable real-time presentation of spectral data in the elution profile, followed by off-line processing by main-frame computer to generate three-dimensional A , A, t surface chromatograms of polynuclear aromatic hydrocarbons. Klatt illustrated the usefulness of flexibility in detection wavelength and demonstrated that isometric projections of oil-sample eluates could be used to characterise different stages of the hydrogenation process. Light 7 1’ Diodes b’ r Electron scan beam Fig. 1. Schematic representation of the diode structure and principles of operation of the silicon vidicon detector (after reference 1).The SIT has also shown promise as a luminescence detector in HPLC. Jadamec et aZ.1° performed repetitive scans of fluorescence emission spectra at 10-s intervals to characterise oil samples during elution by HPLC. This approach has recently been extended to generate the If,&,&,, emission - excitation matrix at points during the elution profile of polynuclear aromatic hydrocarbons, to give highly characteristic qualitative data in four-dimensional space .I1 As an alternative detector in HPLC, the linear diode array (LDA)12 has been vigorously developed since the early work of Dessy et aZ.,13 who demonstrated its capability for rapid capture of the ultraviolet - visible spectrum for metabolic characterisation by automated liquid chromatography. Although the two Reticon 256-element LDA devices in their double- beam detector suffered from the requirement for cooling to -30 “C to obtain a satisfactory SNR, the flexibility of data handling by microcomputer permitted the display of A v c ~ s ~ s t at any A and isometric projection of A , A, t diagrams. The poor sensitivity commented upon by Dessy et a2.was improved by two orders of magnitude by Milano’s gr0up,~,14 who used a similar but uncooled single LDA with ensemble averaging. More recently, the emergence of a commercial ultraviolet - visible spectrophotometer, based on a new LDA design with acceptable SNR at ambient temperature,15 has presented several new avenues for greater specificity and flexibility in HPLC detection.Willis and his group have demonstrated real- time pseudo-isometric displays of A versus A at successive time intervals, illustrated by the separation and characterisation of the three steroids cortisone, dexamethasone and corti- costerone (Fig. 2). Further, when chromatographic bands cannot be efficiently resolved, the commercial LDA detector permits deconvolution of the overlapping bands within 1 s by apply- ing the principle of “over-determination” and least mean squares maximum probability statistics. This technique involves finding the best statistical fit of linear combinations of each component spectrum stored in a reference archive, using all spectral datum points within a specified wavelength interval. It does pre-suppose, however, that each component is known and available in purified form for storagein the spectral archive.The fact that this new facility is available for routine HPLC detection, and does not require external computing support, reflects the remarkable improvement in microcomputer technology during the past few514 ADVANCES I N APPLIED HPLC Anal. Proc. years. In contrast, the SIT described by Klattg relies extensively on external computing facilities to perform least-squares deconvolution and for the A , A, t surface display. 0.146 0.096 0.046 0.004 2 Dexarnethasone '20 240 260 280 300 320 230 250 270 290 310 Wavelengthlnrn Fig. 2. Pseudo-isometric display of ultraviolet spectra during the elution of cortisone, dexamethasone and corticosterone separated by reversed-phase HPLC (after reference 15).In both of these approaches to multi-channel detection in HPLC, the observation wave- length can be programmed with time to optimise detection sensitivity, or to improve selectivity with respect to interfering components whose absorption profiles do not com- pletely overlap the analyte spectrum. Further evidence of identity may be obtained by on-line calculation of absorbance ratios at any specified pair of wavelengths, a feature of relevance in biochemical and forensic analysis, where the extensive reference data available may be stored in data archives for rapid assignation of identity. The principal features of multi-channel detectors, summarised in Table I, and more extensively by Talmi,l have only recently become available in HPLC and will require extensive investigation before it becomes clear whether the additional information is relevant or redundant in practical analysis. TABLE I PRINCIPAL FEATURES OF OPTICAL MULTI-CHANNEL DETECTORS I N LIQUID CHROMATOGRAPHY 1 A veysus t presentation a t any wavelength 2 A vevsus h presentation at any time 3 A , A, t isometric display 4 Variable detection wavelength programmed during elution 5 Absorbance ratios for any pair of wavelengths, programmed for each peak 6 Least mean squares spectral deconvolution of overlapping bands a t any time 7 Luminescence emission excitation matrix a t any time during elution Derivative Techniques in High-performance Liquid Chromatography In contrast with the chemical derivatives frequently generated by pre- and post-column procedures in HPLC, as described recently by Freil6 in an excellent review, the mathematical derivative technique operates solely on the chromatographic detector output to generate the first, second or higher order derivative of the output voltage with respect to a selected variable such as wavelength or time.Higher order derivative spectroscopy has been examined recently by several groups of worker^,^^-^^ who have shown that the even wavelength deriva- tives of Gaussian and Lorentzian bands (d2A/dA2, d4A/dh4, . . .) are often of greater analytical use than the odd functions. This arises from the observation that whereas the odd derivatives are disperse functions, the even derivatives are characterised by a central peak which is narrower than, and coincident with, the zero-order band, The even derivative profile of a mixture displays the constituent bands in sharpened form, as band width decreases with increasing derivative order, and this has led to the wide use of higher derivative spectro- scopy for resolution enhancement .23 Another useful property of the derivative process is that it discriminates against broad bands to an extent which increases with derivative order, 92.The ratio of the amplitudes for two coincident Gaussian or Lorentzian bands 1 and 2 inDecember, 1980 ADVANCES IN APPLIED HPLC 515 the nth derivative, D,, and D,,, is inversely related to the nth power of their band widths W , and W,: DlnlD2n (W,/Wdn This observation, first recorded by Butler and hop kin^,,^ implies that the derivative technique discriminates against broad background absorption (or fluorescence) spectra, and that the relative sensitivity to sharp peaks increases rapidly with derivative order.22 The various mechanisms whereby the derivative method may suppress matrix inerference to reduce systematic error in quantitative spectrometry have been studied recently by computer simulations.22 Although it is now technically possible to generate derivatives of sixth and higher order,3920--23 their use is limited by the appearance of satellite artefacts (two in second, four in fourth order, .. .) extending symmetrically on each side of the centroid peak. These satellites may interfere with adjacent bands, thus making qualitative and quantitative assays more complex.The problem is more acute for Gaussian than for Lorentzian bands, where the prospects for higher order derivatives in resolution enhancement and background correction are much better.,, Progress in applying the derivative method has been noticeable in spectroscopy, although the derivative concepts are of general application in analysis, as illustrated by workers in electrochemi~try,~~ thermal analysis26 and more recently HPLC.,?3 Milano et aL2 were the first to demonstrate that the gradient of the ultraviolet - visible spectrum (dA/dh) could be used to eliminate a component selectively from a chromatogram. This was achieved by setting the derivative observation wavelength of the LDA to coincide with the Amax. (where dA/dA = 0) of the component to be eliminated.Unfortunately, the other components appeared as positive or negative peaks, depending on the numerical value of their first derivative at the observation wavelength. This method may be more selective than the normal single-wavelength detector but suffers from lack of sensitivity, except for those relatively sharp peaks whose maxima occur close to the observation wavelength. A more flexible procedure for resolution enhancement in HPLC, first demonstrated in 1978,3 is to apply the derivative technique to the elution profile itself. The output of the detector, which may be based on the ultraviolet absorbance, fluorescence, refractive index, electrochemical or indeed any linear physical property of analyte concentration, is diff erenti- ated with respect to time to give the second or fourth derivative (d2A/dt2, d4A/dt4). This is readily achieved using an electronic resistance - capacitance device3 or a microcomputer.If the detector signal, Emax., at a peak maximum eluting at time tmax. in the zero-order chromatogram is linearly related to concentration, C , then Emax. = KC at tmax., where K is a constant characteristic of the detector and the chromatographic con- ditions. The nth order derivative with respect to time becomes at tmax.. If a chromatographic band may be described by a Gaussian function defined by E = Emax. exp ( -x2/2) where x = (t - tmax.)/g and 0 is the band half-width at the points of inflection (Le., standard deviation), the second derivative is d2E - _ _ - Emax. (1 - x2) exp (-x”2) dtz - 0 2 and the fourth derivative becomes (3 - 6x2 + x4) exp (--x2/2) d4E Emax.dt4 - u4 --- It is apparent that the sign of the even derivative function at tmax. alternates, being516 ADVANCES I N APPLIED HPLC Anal. Proc. negative in second, positive in fourth, and so on, as illustrated in Fig. 3, where the band width is seen to decrease with increasing derivative order. Another property of the time derivative function is that broad overlapping peaks and base-line variations are progressively eliminated with increasing derivative order. This is depicted in Fig. 4, where the distortion of a Gaussian band superimposed on a strongly curved base line is reduced in each higher even derivative chromatogram. The amplitude measures available for quantitative analysis are also illustrated.The potential benefits conferred by recording the derivative in the time domain apply equally in gas - liquid chromatography (GLC) as in HPLC, and are summarised in Table 11. - Ik i -2 I 1 1 1 Zeroth 1st 2nd 3rd 4th Fig. 3. Zeroth t o fourth derivatives in time domain of a chromatogram of phenol (P, 0.4 mg ml-l). Column, 100 x 5 mm Hypersil, 5 p ~ ; eluent, methanol - water (65 + 35) a t 1.0 ml min-1; pump, LC Series 1 (Perkin-Elmer) ; detector, LC 55-B (Perkin-Elmer) at 272 nm; differentiation units, two Hitachi derivative modules in series (time constant M6) ; syringe injection, 4 p1. As is frequently the case in derivative spectroph~tometry,~~ the SNR of the derivative chromatogram decreases by a factor of approximately two with each higher derivative order, although this depends essentially on the degree of smoothing and the band width of the zero-order peak.27 Hence, one of the early problems experienced in applying the time derivative technique in HPLC was concerned with optimising the SNR.3 Satisfactory performance was obtained with a simple Hitachi differentiation device adjusted to its maxi- mum resistance - capacitance time constant, which incorporates an additional degree of smoothing in the circuit.Provided that the minimum input voltage to the differentiator is kept relatively high (above 50 mV) by selecting the appropriate scale expansion for the detector output, good quality second and fourth derivative chromatograms can be recorded. The higher derivative method has been successfully applied in HPLC for the resolution of overlapping polynuclear aromatic hydrocarbons (Fig.5 ) , paraben preservatives, phenols3 and various drug formulations. Quantitative assays for components resolved in the derivative chromatogram have also been developed. Low-noise microcomputer devices have recently been de~cribed2~9~8 that incorporate refined algorithms for differentiation and smoothing, and with variable smoothing ratios (or time constants) suitable for both GLC and HPLC. Such devices would bring greater sensitivity and extend the scope of this method for chromato- graphic deconvolution. TABLE I1 CHROMATOGRAPHIC FEATURES OF SECOND AND FOURTH DERIVATIVES IN TIME DOMAIN 1 2 3 4 5 Improved resolution of overlapping elution bands Potential increase in sensitivity t o sharp bands Discrimination against interference by broad bands Reduced sensitivity to base-line drift Characteristic elution profile for identification of complex tnixturesDecember, 1980 ADVANCES I N APPLIED HPLC 517 Zero-oraer spectrum Second derivative Fourth derivative of Gaussian band and ’ sloping base-line Q: 40, tR - ----- 2 ----- 0 ’ 1 I Fig.4. Zeroth, second and fourth derivatives of a Gaussian band superimposed on a sloping base line, illustrating the progressive reduc- tion of matrix interference. The various derivative amplitude measures available for quantitation are as follows: Ds, peak-satellite (before t ~ ) ; DL, peak-satellite (after t ~ ) ; DB, peak-tangent base line; D,, peak- derivative base line zero. Derivative Detection with Multi-channel Devices in High-performance Liquid Chromatography Since Milano et aZ.3 work on the use of the first derivative in the wavelength domain for chromatographic deconvolution,2 little consideration appears to have been given to the potential contribution of higher derivative techniques in combination with multi-channel detectors in HPLC. However, now that multi-channel ultraviolet - visible spectrophoto- meters are commercially available equipped with facilities for recording the first and second derivatives with respect to time or wavelength, various new possibilities arise for refining selectivity and improving quantitative reliability in liquid chromatography.First, the derivatives in the wavelength domain could be used to obtain additional evidence of identity for the broad, relatively featureless peaks often encountered in ultraviolet - visible spectro- scopy.An archive of derivative spectra would give many more points of comparison for search- and-find routines in qualitative analysis. The wavelength derivatives of overlapping spectro- scopic bands that co-elute may permit selective quantitative a~sayl’-~O directly. However, when the spectra of overlapping chromatographic components are closely similar, the straight- forward derivative assays break down. In such instances the principle of “over-determination” may be successful, where all the spectral resolution elements are employed in a least mean squares routine. This method may also be applied to the first or higher derivatives of spectral absorbance, an approach which has been shown by several groups of workers to be remarkably effective in deconvoluting overlapping spectral bands and in suppressing broad matrix i n t e r f e r e n ~ e .~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ These approaches to spectral deconvolution do, however, make the 0.6 0.4 0.2 0 0 5 6 7 8 0 5 6 7 8 Time/min Fig. 5. Zeroth and second derivative chromato- grams of phenanthrene and anthracene impurity. Chromatographic conditions as for Fig. 3, but using a Pye Unicam LC-UV detector a t 254 nm.ADVANCES I N APPLIED HPLC Anal. Proc. 518 assumption that the identity of each component is well defined and available in pure form for archival storage, as discussed above, Alternative presentations of data currently employed for qualitative enhancement in HPLC include the A , A, t isometric projections or contour diagrams and the If, Xf, A,, emission - excitation matrix displays discussed above.The replacement of absorbance and fluorescence intensity with their second or higher even derivatives would conceivably give improved resolution between bands. Indeed, the derivative satellite peaks above or below the plane of the derivative base line could be truncated to simplify the display. However, this approach might run the risk of generating redundant data for purposes where a more straightforward approach would suffice. An alternative approach now possible with commercial multi-channel spectrophotometers is to compute the second derivative in the time domain at a fixed detector wavelength. Indeed, the observation wavelength could be specified for each cluster of overlapping bands in the elution chromatogram in order to maximise sensitivity.The use of the multi-channel detector would be more flexible for this purpose than the single-channel detector used in the earlier application of this m e t h ~ d . ~ As the microcomputer derivative function is more likely to reflect the true shape of the derivative chromatogram than that observed with the electronic resistance - capacitance device, better resolution of overlapping bands would be expected by this method. Bearing in mind that second and higher derivative functions are generally unfamiliar in the routine laboratory, it could be useful to integrate the differentiated profile to present the data in more familiar form in those instances where the time derivative technique is used for background correction.Clearly, any resolution enhancement in the derivative display would be sacrified upon re-integration and the SNR would be degraded. However, having identified the relative amplitude and location of the overlapping bands, it would be possible to indicate these parameters in the integrated display, or to apply a con- volution function that would sharpen the bands in the integral chromatogram. Although this discussion has been primarily concerned with the potential contribution of derivative techniques and multi-channel detectors in HPLC, it is clear that the derivative concept is generally applicable to detection in flowing systems. The properties of resolution enhancement and correction for variable base line could be applied to automatic analyser outputs and to the relatively new technique of flow injection analysis (FIA).31 In these systems the problem of peak tailing, which reduces sample throughrut, would be significantly reduced by applying the second or fourth derivative in time domain.The narrower, skewed derivative peaks observed in FIA, for example, are related to concentration in a similar manner to derivative HPLC peaks, and would permit a higher sample injection rate. Preliminary experiments using digital differentiation techniques have confirmed that FIA profiles generated under conventional experimental c0nditions3~ are progressively sharpened in the second and fourth derivatives, where the full widths at half maximum peak height are 4840% and 30-35y0, respectively, of the original zero-order band The potential of derivative techniques in combination with optical multi-channel detectors in liquid chromatography, summarised in Table 111, should not be taken as a substitute for TABLE I11 PRINCIPAL FEATURES OF DERIVATIVE TECHNIQUE COMBINED WITH OPTICAL MULTI-CHANNEL DETECTORS I N LIQUID CHROMATOGRAPHY 1 2 3 4 5 Derivative deconvolution in wavelength domain of co-eluting peaks whose spectra Derivative deconvolution in time domain of overlapping bands in the elution profile Additional evidence of identity by reference to spectral derivative archive Re-integration of differentiated spectra or of time derivative elution profiles to Pseudo-isometric presentation of (derivative function), A, t for improved spectral the classical approach, where all of the relevant chromatographic parameters are systemati- cally optimised to design the most efficient analytical separation.It should be the aim of the chromatographer to apply the minimum of spectroscopic or other refinements as are necessary to attain a given analytical objective. It does seem probable, however, that microcomputer-generated functions will be increasingly available in commercial HPLC overlap eliminate broad band and base-line drift interference resolutionDecember, 1980 ADVANCES I N APPLIED HPLC 519 detectors. Moreover, the new generation of multi-channel ultraviolet - visible and lumines- cence dete~tors,~4 together with the recent emergence of sensitive Fourier-transform infrared detectors,35s36 have added another dimension to detector technology in HPLC.It is the combination of these recent developments with the relatively uncharted areas of derivative chromatography in the time and wavelength domains that presents a creative challenge to the chromatographer interested in extending specificity and selectivity in liquid chromato- Helpful discussions with Dr. B. P. Chadburn, Professor E. Grushka, Professor J. H. Knox and Mr. G. Parsons during the course of this work are acknowledged. The generous loan of Hitachi cliff erentiation modules by Perkin-Elmer Ltd. is also acknowledged. Thanks are extended to Dr. E. H. Hansen, Professor J. RhiiEka and Dr. A. Ramsing for supplying the digitised FIA data. References graphy. 1. 2. 3. 4. 0. 6. 7. 8. 9. 10. 11. 12.13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 36. Talmi, Y ., Editor, “Multichannel Image Detectors,” ACS Symposium Series, No. 102, American Milano, M. J., Lam, S., and Grushka, E., J . Chromatogr., 1976, 125, 315. Fell, A. F., UV Spectrom. Group Bull., 1979, 7, 5. Denton, M. S., DeAngelis, T. P., Yacynych, A. M., Heinemann, W. R., and Gilbert, T. Ur., Anal. Rogers, L. B., Chem. Eng. News, April 15, 1974, 18. McDowell, E., and Pardue, H. L., Anal. Chem., 1976, 48, 1815. McDowell, E., and Pardue, H. L., Anal. Chem., 1977, 49, 1171. Fell, A. F., Anal. Proc., 1980, 17, 266. Klatt, L. N., J . Chromatogr. Sci., 1979, 17, 225. Jadamec, J . R., Saner, W. A., and Saner, R. W., in Talmi, Y., Editor, “Multichannel Image Detectors,” Shelly, D. C., Ilger, W.A., Fogarty, M. P., and Warner, I. M., Altex Chromatogram, 1979, 3, 4. Horlick, G., Appl. Spectrosc., 1976, 30, 113. Dessy, R. E., Nunn, W. G., and Titus, C. A., J . Chromatogr. Sci., 1976, 14, 195. Milano, M. J., Lam, S., Savonis, M., Pautler, D. B., Pav, J. W., and Grushka, E., J . Chromatogr., Willis, B. G., Hewlett-Packard Application Note, No. 8450-5, 1979. Frei, R. W., Proc. Anal. Div. Chem. SOC., 1979, 16, 289. O’Haver, T. C., and Green, G. L., Anal. Chem., 1976, 48, 312. Fell, A. F., PYOC. Anal. Div. Chem. SOC., 1978, 15, 260. Honkawa, T., “Higher Order Derivative Spectrophotometry,” Applications Data Bulletin ADS 112, Talsky, G., Mayring, L., and Kreuzer, H., Angew. Chem., I n t . Ed. Engl.. 1978, 17, 785. Pavlath, A. E., and Millard, M.M., Appl. Spectrosc., 1979, 33, 502. Fell, A. F., UV Spectrom. Group Bull., 1980, 8, 5. Butler, W. L., Methods Enzymol., 1979, 56, 501. Butler, W. L., and IIopkins, D. W., Photochew. Photobiol., 1979, 12, 439. Davis, H. M., and Seaborn, J. E., Electr. Eng., 1953, 25, 314. Erdey, L., Paulik, F., and Paulik, J., Acta Chim. Hung., 1956, 10, 61. O’Haver. T. C., and Begley, T., to be published. O’Haver, T. C., Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Atlantic Hawthorne, A. R., and Thorngate, J. H., Appl. Spectrosc., 1979, 33, 301. Allen, G. C., and McMeeking, R. E., Anal. Chim. Acta, 1978, 103, 73. RbiiEka, J., and Hansen, E. H., Anal. Chim. Acta, 1980, 114, 19. Ramsing, A.. personal communication. Fell, A. F., and Hill, R.G., to be published. Jadamec, J. R., Saner, W. A., and Kallet, E., Pittsburgh Conference on Analytical Chemistry and Vidrine, D. W., J . Chromatogr. Sci., 1979, 17, 477. Malczewski, M. P., and Grushka, E., J . Chromatogr. Sci., in the press. Chemical Society, Washington, D.C., 1979. Chem., 1976, 40, 20. ACS Symposium Series, No. 102, American Chemical Society, Washington, D.C., 1979, p. 115. 1978, 149, 599. Perkin-Elmer, Norwalk, Conn., 1978. City, N. J., 1980, Paper 561. Applied Spectroscopy, Atlantic City, N. J., 1980, Paper 130. Pre-concentration and Selective Sample Handling in High-performance Liquid Chromatography R. W. Frei Department of Analytical Chemistry, The Free University, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands The importance of detection limits and hence the need for sensitivity and selectivity is clearly recognised in trace analytical work.Often chromatographic separation techniques are520 ADVANCES I N APPLIED HPLC Anal. Proc. necessary prior to signal generation, in which event the attainable detection limit can be strongly influenced by chromatographic parameters. Huber et a1.l were among the first to appreciate the problem of achieving low detection limits through the optimisation of chro- matographic operations. More recently, Karger et al.2 have extensively discussed this topic. These and other workers3 also pointed out the importance of using large injection volumes for the improvement of detection limits. This is particularly valid when conventional procedures for sample concentration become difficult owing to solubility problems or the instability of the samples.The model studied by Karger et aL2 did not consider the situation of solutes that are strongly retained on the top of the column under injection conditions. In many trace analytical procedures, particularly in chromatography, such a true trace- enrichment (or pre-concentration) step is unavoidable prior to quantitative analysis to attain the necessary concentration of compounds for the detection step. Classical techniques such as freeze-drying, extraction and/or evaporation, steam distillation, etc., have serious limitations in terms of recovery, capacity and possible loss of sample during processing, especially for temperature-sensitive compounds. Trace enrichment via adsorption on solid surfaces has therefore been adopted in conjunction with gas-chromatographic techniques.A well known example is the carbon adsorption method (CAM), recently reviewed by Suffet and S~winski.~ Grob5 has developed a closed-circ nit carbon adsorption system, which has been widely used for the trace enrichment of organics from aqueous samples. Hydrophobic- layer materials have also been introduced for relatively low-polarity organics. Junk et aZ.6 have reported excellent recoveries (90-100%) for compounds such as pesticides, alkyl- benzenes, halogenated compounds, polynuclear aromatics and phenols, using a macroreticular XAD-2 resin packed into glass tubes for sampling of litre volumes of water samples. Other materials such as XAD-4 resin and Spherocarb were discussed by Tateda and FritzT In recent years the use of high-performance liquid chromatographic (HPLC) packing materials for trace enrichment either on-column or on-line with pre-columns has become a further powerful tool in trace analytical work.Both non-polar and polar materials, almost invariably based on a silica gel matrix, have been used. Hydrophobic surfaces, such as are present on the commercially available non-polar C, or C,, reversed-phase materials, lend themselves readily to the collection of a wide range of organics. In 1974, Kirkland8 discussed the possibility of using pre-concentration phenomena on the top of analytical HPLC columns for the purpose of trace enrichment. Little and Fallick9 reported on the use of very large injection volumes of up to 200ml for relatively non-polar compounds applied from an aqueous solution to a C,, reversed-phase column.Similar results were reported for peptides and alkaloids injected from aqueous solutions.10-12 When relatively non-polar organic species are injected from an aqueous solution on to a hydrophobic surface they will become immobilised until the elution strength of the solvent mixture is increased. This means that the compounds are concentrated into a very small zone on top of a reversed-phase chromatographic column (trace-enrichment effect). The components can then be eluted with a suitable agent with very little band broadening. Similar phenomena have been observed in adsorption systems for relatively polar compounds injected from non-polar solvents. l3 This method is thus well suited for the direct injection of, for example, aqueous pharma- ceutical preparations, biological fluids and, particularly, aqueous samples in connection with water pollution studies.This has been demonstrated by Creed,14 who analysed a number of samples such as industrial effluent water, polluted river water and well water. Otsuki15 successfully studied the trace enrichment of phthalate esters, which are widely used as plasticisers, from aqueous samples on a VBondapak C,, column, with subsequent elution by means of a water - methanol gradient. Good reproducibility of repetitive large-volume injections can be obtained using a timer-controlled step-gradient apparatus,16 which can also be used to carry out a gradient separation of the pre-concentrated compounds.This can be seen in Fig. 1, showing the isolation of about 2 p.p.m. of an ergot alkaloid from a urine sample injected without preliminary clean-up. Pre-column Techniques The above approach is useful for trace analysis in complex matrices. The direct injection of large amounts of sample and/or very dirty samples on to anDecember, 1980 ADVANCES IN APPLIED HPLC 52 1 analytical column can exert much strain on this column. It is frequently observed that after a few injections of, for example, biological fluids or their extracts the separation performance of the column decreases drastically. HPLC columns are expensive and high- efficiency columns are valuable. The use of short pre-columns has therefore been recom- mended in order to protect the analytical column17 and, simultaneously, to provide a partial clean-up of the sample.In addition, the pre-column can be used efficiently in the trace- enrichment step and can possibly be employed for field-sampling methods and even for on-column derivatisation. Application of the latter possibility has been demonstrated by Maitra et aZ.lB for the off-line derivatisation of gentamicin in human serum. An interesting application of the pre-column concept has been proposed by Lankelma and Poppel9 for the determination of the cytostatic drug methotrexate in plasma. Huber and Becker13 have recently recommended the use of so-called displacement HPLC for trace enrichment. Polar compounds are pre-concentrated on a silica gel pre-column, while reversed-phase materials are used with hydrophobic model compounds.Using a displace- ment solvent of suitable polarity, high enrichment factors of typically between about 1000 and 5000 are obtained. For the trace-enrichment of phthalate esters from aqueous samples, Ishii et aZ.20 used miniaturised HPLC equipment .21 This miniaturised version is also suitable for bioanalytical applications with corticosteroid analysis in serum samples.22 However, the real value of the pre-column approach is in an on-line mode, which can be automated for routine analysis of large series of samples. Such an approach has, for example, been discussed by May et aZ.,23 who reported on the analysis of hydrocarbons in marine sediments and sea water. Eisenbeiss et aZ.24 developed a special adsorbent for the enrichment step, which allowed them to elute and separate pre-concentrated polycyclic aromatic hydrocarbons under iso- cratic conditions.In all of the above instances relatively long pre-columns packed with small-diameter particles have been used. In all probability, this will often be unnecessary as solutes displaying high capacity factors under trace-enrichment conditions will be pre-concentrated on a thin top layer of the pre-column. Long columns, moreover, have the disadvantage of a slow sample throughput owing to a high back-pressure. an attempt was therefore made to work with very short pre-columns having a sorbent layer thickness of only 2 mm. Fig. 2 shows the design of the pre-column, which is hand-packed with a dense The above procedures are based on the off-line pre-column concept. In a recent DHCM - + 40% Start H20 60%CHICN H20 1 L I 15 10 5 0 Time/m in 1 2 3 4 5 6 Fig.2. Pre-column design. 1, Swagelok, t x Q in; 2, varying- length PTFE rod, 4.6 mm 0.d. ; 3, stainless-steel tube, 45 x 4.6 mm i.d.; 4, stainless-steel capillary, 6 in o.d., 0.25 mm i.d.; 5, PTFE or stainless-steel 20-pm frit; 6, Swagelok, + x Q in, containing a 0.5- or 2-pm outlet frit. The space between 5 and the outlet frit of 6 is filled with packing material.25 Fig. 1. Chromatogram of 2.74 p.p.m. of dihydroergocristine mesilate (DHCM) in urine.12522 ADVANCES I N APPLIED HPLC Anal. Proc. slurry of the stationary phase material in methanol using a micro-spatula. Phthalate esters are used as model compounds, and samples of up to 1 1 of, e.g., tap water, demineralised water and mineral water typically containing 0.1-2 parts per 1 0 9 (p.p.b.) of phthalate esters are pre-concentrated on 5-pm LiChrosorb RP-18 with 95-100% solute recovery, using pumping speeds of 5-25 ml min-l.The analytical column is also packed with LiChrosorb RP-18, and methanol - water mixtures are used for elution. A typical high-performance liquid chromatogram recorded for a sample of mineral water [found to contain 1.2 p.p.b. of di(ethylhexy1) phthalate and 0.08 p.p.b. of dibutyl phthalate] is shown in Fig. 3. 0 8 1 6 0 10 20 30 Retention tirnehin Fig. 3. (a) High-performance liquid chromatogram recorded for. a standard solution (- - -) of dibutyl and di(ethylhexy1) phthalate, and for a sample (-) of mineral water. Conditions : sample solution, 800 ml; 2 x 4.6 mm 5-pm LiChrosorb RP-18 pre-column; loading a t 25 ml min-l pump- ing speed; analysis a t 2 ml min-l on a 12.5 cm x 4.6 mm i.d.LiChrosorb RP-18 separation column with 75% (60 s), 85% (300 s) and 95% (270 s) methanol in water as mobile phase; detection a t 233 nm (0.08 A.U.F.S.).S5 Comparison of high-performance liquid chromatograms recorded for equal amounts (about 20 pg) of the PCB mixture Aroclor 1260, using (b) normal 200-pl syringe injection, or (c) trace enrichment from a 16.5-ml aqueous sample. Mobile phase, 85% methanol in water; detection at 205 nm. For pre-column and separation column, see previous c ~ n d i t i o n s . ~ ~ The principle outlined above has also been used su~cessfully~~ for the trace enrichment of chloroanilines and polychlorinated biphenyls (PCBs) .Fig. 3 illustrates well the negligible contribution of the short pre-column to band broadening, using a complicated PCB mixture as a model “compound.” Further Trends and Future Developments One of the inherent disadvantages of trace enrichment on surfaces of low selectivity is that many sample constituents will be concentrated on the pre-column. Consequently, even if overloading problems are neglected, clean-up may well be insufficient and HPLC separation of the compound of interest will be rendered difficult. There are two chief routes to eliminate this problem, vix., via an increase of selectivity effected by post-column (and also pre- or on-column) derivative formation, or via multi-dimensional HPLC. An example of on-column derivatisation, vix., the analysis of gentamicin as its o-phthalde- hyde derivative,ls has already been cited ; a post-column reaction between peptides and fluorescamine was used in earlier work.l1,l7 The combined use of trace-enrichment and post- column derivatisation has also been applied successfully in the determination of chloro- pheniramine in urine.26 This tertiary amine drug was pre-concentrated from 2 ml of a 1 : 20 diluted untreated urine sample on a LiChrosorb RP-18 pre-column.Subsequently, analysisDecember, 1980 ADVANCES I N APPLIED HPLC 523 was effected on a Micropak CN-10 column using methanol- water (1 + 4) containing phosphate buffer as the mobile phase. Detection was based on ion-pair formation between the amine cation and the fluorogenic dimethoxyanthracene sulphonate (DAS) and extraction of the ion pair into tetrachloroethane in a micro-extraction reactor, monitoring (part of) the organic solvent stream in a fluorimeter.In a related study2' (see Fig. 4) hydroxyatrazine (HA), which is the major metabolite of the well known herbicide atrazine, was taken as the model compound. With this relatively polar compound, pre-concentration from 10-ml samples on a 2-mm LiChrosorb RP-18 layer was found to be critical with regard to recovery owing to breakthrough of HA. Reducing the polarity by ion-pair formation with DAS and trace enrichment of HA as its HA - DAS ion pair effectively remedied this situation. Time d Fig. 4. Chromatogram for hydroxyatrazine (H) in urine after pre- concentration. I.Blank urine; and 11, urine spiked with 0.5 p.p.m. of HA. 10ml of urine pre-concentrated each time. Chromatographic conditions : RP-2 column and a mobile phase consisting of 25% methanol - 0.1 N sodium dihydrogen orthophosphate solution, pH 3.5; flow-rate, 1.0 ml min-l. (a) UV trace a t 225 mm; and (b) fluorescence trace. An alternative means of solving the selectivity problem is the multi-column a p p r o a ~ h ~ ~ ~ ~ with on-line coupling of one column system to another. In order for the transfer of zones to occur with a minimum of band broadening it is necessary to have a trace-enrichment step on the second, and subsequent, column(s). Future research will no doubt also be directed to the use of more selective pre-column packing materials, such as metal-loaded solid surfaces and immobilised enzyme-cont aining phases.Different pre-columns may be used in series to isolate different groups of compounds for final analysis. Attention should further be focused on the use of pre-columns for field sampling. In this way, the samples can be considerably reduced in size and stabilised for storage25 and transportation to a central laboratory. In our opinion, the interest of analytical chemists in these and other trace-enrichment techniques will certainly be stimulated by the automation potential of the pre-column and the multi-dimensional HPLC concept. References 1. 2 . Huber, J . F. K., Hulsman, J. A. R. J., and Meyers, C. A. M., J . Chromatogr., 1971, 62, 79. Karger, B. L., Martin, M., and Guiochon, G., Anal. Chem., 1974, 46, 1640.524 3.4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. ADVANCES IN APPLIED HPLC Anal. Proc. Strubert, W., Chromatographia, 1973, 6, 205. Suffet, I., and Sowinski, E. J., Jr., in Grob, R. L., Editor, “Chromatographic Analysis of the Environ- ment,” Marcel Dekker, New York, 1975, p. 435. Grob, K., J . Chromatogr., 1973, 84, 255. Junk, G. A, Richard, J . J., Fritz, J . S., and Svec, H. J., in Keith, L. H., Editor, “Identification and Analysis of Organic Pollutants in Water,” Ann Arbor Science Publishers, Ann Arbor, Mich., USA, 1976, p. 135. Tateda, A., and Fritz, J . S., J . Chromatogr., 1978, 152, 329. Kirkland, J , Analyst, 1974, 99, 859. Little, J . N., and Fallick, G. J., J . Chromatogr., 1975, 112, 389.Krummen, K., and Frei, R. W., J . Chromatogr., 1977, 132, 27. Krummen, K., and Frei, R. W., J . Chromatogr., 1977, 132, 429. Schauwecker, P., Frei, K. W., and Erni, F., J . Chromatogr., 1077, 136, 63. Huber, J . F. K., and Recker, R. R., J . Chromatogr., 1977, 142, 765. Creed, C. G., Res./Dev., September, 1976, 40. Otsuki, A., J . Chromatogr., 1977, 133, 402. Erni, F., Frei, R. W., and Lindner, W., J . Chromatogr., 1976, 125, 265. Frei, R. W., Int. J . Environ. Anal. Chem., 1978, 5, 143. Maitra, S. K., Yoshikawa, T. T., Hansen, J. L., Nilsson-Ehle, I., Palin, W. J., Schotz, M. C., and Lankelma, J., and Poppe, H., J . Chromatogr., 1978, 149, 587. Ishii, D., Hibi, K., Asai, K., and Nagaya, M., J . Chromatogr., 1978, 152, 341. Ishii, D., Asai, K., Hibi, K., Jonokuchi, T., and Nagaya, M., J , Chromatogr., 1977, 144, 137.Ishii, D., Hibi, K., Asai, K., Nagaya, M., Mochizuki, K., and Mochida, Y., J . Chromatogr., 19’78, May, W. E., Chester, S. N., Cram, S. P., Cump, B. H., Hertz, H. S., Enagonio, D. P., and Dyszel, Eisenbeiss, F., Hein, H., Joester, R., and Naundorf, G., Chromatogr. N e d . , 1978, 6, 8. Brinkman, U. A. Th., van Vliet, H. P. M., and Frei, R. W., J . Chromatogr., 1979, 185, 483. Frei, K. W., Lawrence, J . F., Brinkman, U. A. Th., and Honigberg, I. L., J . High Resolut. Chromatogr. van Buuren, C.. Lawrence, J . F., Brinkman, U. A. Th., Honigberg, I. L., and Frei, R. W., Anal. Chem., Erni, F., and Frei, R. W., J . Chromatogr., 1978, 149, 561. Johnson, E. L., Gloor, R., and Majors, R. E., J . Chromatogr., 1978, 149, 571.Gruze, L. B., Clin. Chem., 1977, 23, 2275. 156, 173. S . M., J . Chromatogr. Sci., 1975, 13, 535. Chromatogr. Comnzun., 1979, 2, 11. 1980, 52, 700. Mechanism of lon-pair Reversed-phase High-performance Liquid Chromatography Richard A. Hartwick and John H. Knox” Department of Chemistry, University of Edinburgh, West Mains Road, Edinburgh, EH9 3 J J Considerable ambiguity and controversy has arisen over the precise chemical mechanism (s) by which ion-pairing agents effect the retention of ionised solute molecules when using reversed-phase, chemically bonded packing materials. Some workers1 have felt that the primary mechanism involved is the formation of ion pairs in the mobile phase, with the enhanced retention of the ion-pair complex being due to increased hydrophobic interactions. In this theory, the solute need not be present exclusively as ion pairs in the eluting agent but rather an equilibrium is thought to exist between the solute ion and the solute ion pair; the two species are then partitioned into the stationary phase to different extents.An alterna- tive view is that the stationary phase is first coated with the pairing ion present in the mobile phase, and the adsorbed ionogens create a kind of dynamic ion e ~ c h a n g e r . ~ , ~ Obviously, neither mechanism needs to function exclusively. The exact mechanism must be some intermediate between the two, depending on the pairing agent used and the nature of the chromatographic systems. Through observations of the chromatographic behaviour of ionised solutes on reversed- phase, ion-pair systems, we were led to the idea that the second mechanism, i.e., the ion- exchange mechanism, was predominant when using moderately long-chain pairing agents and normal elution conditions.The basic premise of this research was that, as a first approxi- mation, retention ought to be dependent solely on the surface charge density of the column packing resulting from the adsorption of pairing agent molecules on to the stationary phase. * To whom correspondence should be addressed.December, 1980 ADVANCES IN APPLIED HPLC 525 Hence, it was felt that retention ought to be primarily dependent on the number of ionogens per unit area of the packing rather than on the chain length or concentration of the pairing ions per se. Accordingly, we have measured the adsorption isotherms of a number of long-chain sulphates (C8, C,,, C12) on ODS Hypersil (a 5 pm maximally capped octadecylsilica of about 200m2g-l surface area) using the breakthrough method; we have also determined the k’ values of positively charged, neutral, negatively charged and zwitterionic species on columns equilibrated with these surface-active agents.In order to examine our basic hypothesis the values of k’ have been plotted against the surface concentration of surface-active agent. Some typical results are shown in Fig. 1. The excellent linear relationship observed over a substantial concentration range confirms the validity of the general idea. Although the chain length of the surface-active agent has a small effect, the predominant factor governing retention is undoubtedly the magnitude of the surface charge density.25 20 15 3r 10 5 0 0.6 1.2 1.8 2.4 3.0 Surface concentration/fimol m-* Fig. 1. Surface charge in ion-pair chromato- graphy. Packing, ODS Hypersil. Pairing ion, decyl sulphate. A, TYR-AMD+; B, METADR+; C, ADR+; D, PHE-GLY-GLY; E. phenol; F, TRP; and G, naphthylamine sulphonate-. The adsorption of charged species on to the surface of the packing material also gives rise to an electrical double layer and can be interpreted in a quantitative manner in terms of the Stern - Gouy - Chapman (SGC) theory of the double layer, as discussed by Cantwell and P u ~ n . ~ This model provides a qualitative theoretical framework not only for retention through formation of ion pairs but also for exclusion of co-ions, which is clearly demonstrated by the data in Fig.1. References 1. 2. 3. Horvath, C., Melander, W., Molnar, I., and Molnar, P., Anal. Chem., 1977, 49, 2231. Knox, J . H., and Laird, G. R., J . Chromatogr., 1976, 122, 17. Konijnendijk, A. P., and van de Venne, J . L. M., in Zlatkis, A., Editor, “hdvances in Chromato- graphy 1979, Proceedings of 14th International Symposium, Lausanne, Switzerland,” Chromato- graphy Symposium, Department of Chemistry, University of Houston, Texas, USA, 1979, p. 451. 4. Cantwell, F. F., and Puon, S., Anal. Chem.. 1979, 51, 623.526 ADVANCES I N APPLIED HPLC Anal. Proc. Analytical and Preparative H igh-performance Liquid Chromatography of Polypeptide and Protein Hormones from Normal and Pathological Calcitonin-containing Samples E.C. Nice and M. J. O’Hare Ludwig Institute for Cancer Research, Royal Marsden Hospital, Sutton, Surrey, SM2 5PX Establishing the functional characteristics of endocrine and paraendocrine tumours may be of considerable value in both clinical and experimental studies of neoplasia. Although direct radioimmunoassay (RIA) is used routinely for the measurement of hormonal poly- peptides and proteins, a number of factors may influence the values that are obtained: non-hormonal serum components may give anomalous results, there is often considerable variation in the specificity of individual antibodies used for assay and circulating levels of certain hormones (e.g., calcitonin) may be very low, requiring concentration of large volumes of sample for accurate determination and further characterisation.In addition, there is increasing awareness of the existence of large relative molecular mass forms and/or pre- cursors of some polypeptide hormones. These may have different affinities for an antibody (immunochemical heterogeneity), a factor that complicates interpretation of RIA data. We have shown that high-performance liquid chromatography (HPLC) can be used to separate and simultaneously to measure a large number of physiologically important steroids1 in biological samples, enhancing the specificity of RIA and thereby allowing the rapid evaluation of the functional attributes of both normal and neoplastic steroidogenic tissues in vitro. We have, therefore, recently explored the potential of HPLC techniques in an analogous study of polypeptide and protein hormones, including calcitonin and its pre- cursors.Materials and Methods The separation of polypeptides and proteins has been achieved in this study using hydro- phobic interaction chromatography on alkylsilane bonded (reversed-phase) stationary phases. For reproducible high-efficiency chromatography with high recoveries of these compounds in complex mixtures it was necessary to use a salt-containing aqueous primary solvent of low pH (< 4) and high molarity (> 0.1 M) with gradient elution employing a secondary organic solvent (e.g., acetonitrile). Full details of the chromatographic systems and sources of materials have been described Biological samples (e.g., serum, tissue culture medium and cell homogenates) are processed as follows.Trace Enrichment by HPLC “Mini-column” Soluble preparations from the cells, or their supernatant medium, are acidified with hydrochloric acid (pH 2.1) after the addition of pepstatin A, a proteolysis inhibitor, and trace amounts of [3H]-substance P, undecapeptide, are added as an internal recovery and chromatography standard. Samples are then pumped directly on to an 80 x 4.6 mm i.d. stainless-steel column tap-packed with Partisil 10 ODS (pore size 5.5-6.0 nm). The column is washed with 20% acetonitrile - 0.155 M sodium chloride of pH 2.1 (5 ml) prior to elution with 50% acetontrile - 0.155 M sodium chloride of pH 2.1 (5 ml). This procedure recovers most of the immunoreactive calcitonin (iCT) from the column in the latter fraction, from which the acetonitrile is then evaporated under nitrogen, and the detailed chromatographic profile of the hormone(s) and their relative retention times are established by analytical/ preparative HPLC.Analytical/Preparative HPLC Mini-column fractions are chromatographed on a 100 x 5 mm i.d. stainless-steel column slurry packed with a high-coverage 5-pm ODs-silica (e.g., ODs-Hypersil or Nucleosil 5 Cle). Peptides and proteins are eluted using a multi-step linear gradient of acetonitrile in acidic saline (pH 2.1,0.155 M) (see Fig. l), the elution profile being monitored by ultraviolet absorption (225 nm) and endogenous tryptophan fluorescence (275 nm activation). Aliquots of the eluate are then trapped (1-ml fractions) in tubes containing pepstatin A (5 pg per tube) andDecember, 1980 ADVANCES IN APPLIED HPLC 527 samples taken for specific detection of calcitonin-like materials by RIA and for subsequent relative molecular mass determination.The recovery and chromatographic profile of the [3H]-substance P internal standard is established by liquid scintillation counting. 12, 10 8 I - E F, 6 --. 4 4 L-Trp * 0 CTMSO 1 [3H],S~bP ,’ ACTHI-24 4 ,/ J c 0 10 20 30 40 50 60 70 Retention time/min Fig. 1. Hydrophobic HPLC of 1 ml of normal human serum to which 10 ng of calcitonin monomer (CT) had been added. Analytical fractionation was carried out after mini-column trace enrichment as described. The separation was performed a t 35 “C on Nucleosil 5 C18 (100 X 5 mm id.) with a primary solvent of 0.155 M sodium chloride solution (pH 2.1) and acetonitrile as the secondary solvent.The dotted line shows the gradient profile. The lower solid trace is the UV absorption a t 225 nm (1 a.u.f .s.). Tryptophan (10 p g of ~-Trp) and aACTH,-,, (25 pg) were added prior to injection as “visible” chromatography standards. The position of the [3H]- substance P is also shown. CT and CTMSO indicate the retention times of human calcitonin monomer and its methionine sulphoxide determined in a subsequent calibration run. The inset indicates the recovery of iCT in l-ml eluate fractions. Results and Discussion Having optimised the chromatographic conditions for the separation of a number of polypeptides and smaller proteins, and in an attempt to clarify the production of “marker” hormones by some human tumours, we investigated the ectopic production of iCT by a cultured human lung tumour cell line (BEN).5 In order to detect the small amounts of hormone produced in this system (nanograms per lo6 cells per day) a trace-enrichment method was devised.Using the simple mini-column technique described above, recoveries of over SOY-, of iCT, substantially purified from large proteins also present in cell extracts or tissue culture medium, were obtained from samples containing up to 100 mg of total protein (e.g., 100 ml of medium). The characteristic profiles of iCT which were then established by analytical/preparative gradient elution chromatography have been illustrated el~ewhere.~ This system is also capable of handling at least 10 mg of peptides or proteins without loss of resolution.When pepstatin A was included throughout the extraction - purification procedures a late-eluting major iCT component of relative molecular mass 40 kD was detected in both the528 ADVANCES IN APPLIED HPLC Anal. Proc. tissue culture medium and cell extracts,6 illustrating the capability of these hydrophobic interaction systems to separate and purify at least some relatively large materials. Less than 10% of iCT from these cells chromatographed as calcitonin monomer (3.5 kD). In order to compare these results with calcitonin synthesis and secretion in vivo we have examined the behaviour of corresponding materials in serum and whole tissue samples. Neat serum, or soluble homogenates from normal thyroid tissue, are too viscous for direct application to the HPLC mini-column. However, by dilution of the serum with acidic saline the equivalent of 8 ml can be processed before excessive column back-pressure develops.With the thyroid, preliminary removal of thyroglobulin (600kD), by a method such as density centrifugation, is necessary if such samples are to be processed by HPLC for trace enrichment of calcitonin(s). The successful application of these methods to serum samples is illustrated in Fig. 1 . A l-ml volume of normal serum (iCT < 100 pg) was spiked with 10 ng of synthetic calci- tonin monomer. After trace enrichment and analytical HPLC more than 90% of radioimmunoassayable material was recovered with a retention time identical to that of the calcitonin monomer standard chromatographed alone.The presence of large amounts of non-hormonal material in the extracts did not, therefore, significantly modify the chromatographic behaviour of the polypeptide (see the ultraviolet absorption profile in Fig. 1). Only a small proportion of the iCT was found as the earlier eluting methionine sulphoxide form. In preliminary experiments on serum from a patient with neoplasia of the calcitonin-producing cells of the thyroid (medullary thyroid carcinoma) two major peaks of iCT were demonstrated, one late-eluting component and the other corresponding to human calcitonin monomer. We are grateful to Mr. M. Capp for excellent technical assistance and to Drs. M. Ellison and J. Lumsden for helpful discussions and the gift of cell cultures. References 1. 2. 3.4. 5. 6. O'Hare, 31. J., Nice, E. C., Magee-Brown, R., and Bullman, H., J . Chromatogr., 1976, 125, 357. Nice, E. C., and O'Hare, M. J., J . Chromatogr., 1979, 162, 401. O'Hare, M. J., and Nice, E. C., J . Chronzatogr., 1979, 171, 209. Nice, E. C., Capp, M., and O'Hare, M . J., J . Chromatogr., 1979, 185, 413. Ellison, M., Woodhouse, D., Hillyard, C., Dowsett, M., Coombes, R. C., Gilby, I<. D., Greenberg, Lumsden, J., Ham, J., and Ellison, M. L., Biocheni. J . , in the press. P. B., and Neville, A. hl., Brit. J . Cancer, 1975, 32, 373. Extra-thermodynamic Relationships in lon-pair High-performance Liquid - Solid Chromatography C. M. Riley and E. Tomlinson" School of Pharmacy and Pharnaacology, University of Bath, Claverton Down, Bath, B A 2 7 A Y , and Sub- faculty of Pharmacy, University of Amsterdam, Plantage Muidergracht 24, Amsterdam, The Netherlands The analytical armoury provided by modern liquid chromatography can be augmented by ion-pair techniques,l particularly those involving the use of surfactants as pairing ions.As demonstrated by many studies, the latter approach can be advantageous for the resolution of ionised solutes (protic and aprotic), due to the high flexibility and reasonable efficiency afforded. In particular, the technique has uses for the analysis of drugs and related com- pounds in complex formulations and biological fluids. As part of a continuing studyz-6 examining the behaviour of functional groups in liquid chromatography, we have examined the influence of organic solvents and temperature on group retention in ion-pair reversed-phase high-performance liquid - solid chromatography (RP-HPLSC).Our findings are surveyed briefly in this paper, and show how frequently it is possible to use linear free-energy relationships7 to describe and predict retention. * To whom correspondence should be addressed.December, 1980 ADVANCES I N APPLIED HPLC 529 Effect of the Nature and Concentration of Mobile Phase Organic Modifier Schoenmakers et aZ.8 have predicted from theoretical models that the capacity ratio, K, of a solute will vary quadratically with the volume fraction, #, of organic modifier in a mobile phase binary mixture, i.e., where A , B and C are regression coefficients. Equation (1) should be useful in describing not only the effect of organic solvents per se on retention but also in determining optimum gradient profile^.^,^ It has been suggestedg that all solute retention data should be presented in terms of equation (2), which represents a linear relationship between log K and 4, i.e., 10gK = D+ + E where D and E are regression coefficients.The effects of increasing the concentrations of four organic modifiers, i.e., methanol, acetonitrile, propan-2-01 and tetrahydrofuran, on the retention of a number of substituted benzoic acids in their ionised forms using a constant low concentration of alkylbenzyldimet hylammonium chlorides1* as pairing ions are given in Table I according to equation (2). In all instances the solute retention decreased with increase in organic modifier concentration. Only for polar solutes in tetrahydrofuran could equation (1) be used to give a better fit of the data. Values of D and E (Table I) are dependent on substituent character, with D values reflecting the effect of organic modifier on solute retention.The highest values of D are thus obtained with the modifier that has the greatest extracting ability for solute ion pairs.5 Hence, based on the values for benzoic acid, these extracting efficiencies are propan-2-01 (- 10.9) > tetrahydrofuran (-9.3) > acetonitrile (-7.9) > methanol (-4.7). Clearly, such values could form the basis of an elutropic series for solvents used in ion-pair RP-HPLSC. TABLE I REGRESSION COEFFICIENTS FOR EQUATION (2) (EXAMPLES) Benzoic acid substituent H 2-NH2 4-NH2 2-OH 4-OH 2-CH3 4-CH3 Methanol -7 D E -4.74 2.94 -5.22 3.10 -3.21 1.43 -6.18 4.02 -4.22 2.20 -4.71 3.02 -5.35 3.53 Propan-2-01 & D E -10.9 3.39 -11.2 3.38 -6.23 1.43 - - -9.18 2.45 -10.2 3.19 -12.3 4.05 Acetonitrile r-- D E -7.94 2.80 -8.18 2.88 -4.54 1.24 - - -5.27 1.58 -7.89 2.79 -9.38 3.49 Tetrahydrofuran D E -9.29 2.96 -9.73 3.08 -6.46 1.44 -7.39 2.12 -8.60 2.73 - - - - The effect of structure on the values of D can be seen by using a group contribution term AD, such that where j and i refer to substituted and unsubstituted benzoic acids, respectively.AE can be derived from Table I and it can be seen that AD = D, - Di (3) Similarly, T = AD+ + AE (4) where r is an extra-thermodynamic parameter describing the effect that a group has on retention.5 Relating AD and AE to other (non-chromatographic) functional group terms, we can write, for example, for methanol n = 10; y2 = 0.85 (5) AD = -1.117~ - 0.36 AD = -0.98~ - 0.560 - 0.22 n = 10; R2 = 0.93 (6) AE = 1 .1 3 ~ + 0.23 n = 10; r2 = 0.91 AE = 1 . 0 4 ~ + 0.510 - 0.12 n = 10; R2 = 0.98530 ADVANCES I N APPLIED HPLC Anal. Proc. where w and (T are the Hansch hydrophobic termll and the Hammett electronic term,l2 respectively, and n, Y and R refer to the number of data points, the correlation coefficient and the multiple correlation coefficient, respectively. These typical results illustrate that both the slope and intercept terms are strongly dependent on the hydrophobicity term, although electronic effects are significant. (Interestingly, the intercept term, AE, has some electronic contribution, because AE = 7 when 4 --f 0, although we have shown5 that when methanol is present the correlation of T with T is not improved by incorporation of the 0 term.) From a practical viewpoint these data show that D will become increasingly more negative with increasing solute hydrophobicity.13 Organic Solvent Time Normalisation Equations (1) and (2) show how it is possible to achieve a required retention for any solute using any particular organic modifier.To compare functional group selectivity we have used the method of organic solvent norma1i~ation.l~ Thus, by choosing concentrations of organic modifiers to give the same retention of a reference solute, benzoic acid, in methanol (# = 0.50), differences in selectivity can be obtained from equation (9), which relates substitu- ent values obtained using methanol, 7, to those obtained, 7*, in “equivalent” mobile phases of the other modifiers, i.e., r = Fr* + G The regression and correlation coefficients obtained in the present study, for this expression, are given in Table 11.Although propan-2-01 behaves similarly to methanol, the relation- ships for the other modifiers are perturbed by the effect of the polar functions (NO,, NH,, OH), although hydrophobic group selectivity is shown to be independent of the organic modifier, as shown in Table 111. (9) TABLE I1 REGRESSION COEFFICIENTS FOR THE RELATIONSHIP GIVEN BY EQUATION (9) Modifier F G Y n Acetonitrile . . . . 1.03 -0.09 0.98 15 Tetrahydrofuran . . . . 0.85 0.07 0.92 15 Propan-2-01 . . . . 0.99 - 0.01 0.99 15 The perturbances of the relationships embodied in equation (9) are more apparent at low modifier concentrations and their manifestations can be exemplified by Table IV, which gives the capacity ratios of some polar substituted benzoic acids in “equivalent” concentra- tions of propan-2-01 and tetrahydrofuran.These data indicate that separations may be improved by changing to an “equivalent” concentration of different modifier, if the solute polarity is different. Scheme I represents the “equivalent” concentrations for the four modifiers studied here. A 0.20 0.40 0.60 0.80 C 1 0.20 0.25 0.30 ?* 0.20 0.25 0.30 - B 0.20 0.25 0.30 Scheme 1. Equivalent concentrations of organic modifiers (volume fraction). A = Methanol; B = propan-2-01; C = acetonitrile; and D = tetrahydro- furan.December, 1980 ADVANCES IN APPLIED HPLC 53 1 TABLE I11 EFFECT OF ORGANIC MODIFIER ON T Modifier 4 TCH3 TcI Methanol .. . . . . 0.50 0.30 0.53 Acetonitrile . . . . 0.275 0.31 0.57 Propan-2-01 . . . . 0.255 0.31 0.54 Tetrahydrofuran . . . . 0.265 0.28 0.57 Relationship Between T and T Group contribution values obtained from chromatographic systems (7) can be well correlated with terms such as n obtained from non-chromatographic systems. We have shown5 that equation (10) is highly significant in the correlation of T values (water - octan- 1-01) with T values obtained using ion-pair RP-HPLSC with methanolic aqueous mobile phases and various pairing ions : T = 0 . 4 7 ~ - 0.002 (fi = 44; Y = 0.941) (10) By analysing the data from the present study in terms of the general free-energy relationship for equation (lo), i.e., T = ~ T + r (11) it can be seen from Fig.1 that the slope coefficient, H , decreases with increasing value of 4. For the two alcohols the latter relationship is found to be linear and is curved for acetonitrile and t e t rah ydrof uran. These results indicate that bulk-phase partition coefficients (which are available from the literature) can be useful for the prediction of retention and selectivity in ion-pair RP-HPLSC but that equations such as equation (11) would have to be modified (perhaps to include an electronic term) before they could be used for predictive purposes. TABLE IV CAPACITY RATIOS FOR SUBSTITUTED BENZOIC ACIDS UNDER IDENTICAL CONDITIONS USING EQUIVALENT CONCENTRATIONS OF PROPAN-2-OL (4 = 0.20) AND TETRAHYDROFURAN (4 = 0 .2 0 ) Benzoic acid r substituent Propan-2-01 4-NH2 1.4 4-OH 3.1 3-OH 10.6 2-NO, 10.6 H 18.2 3-NH2 2.5 2-NH2 15.0 K -A -i Tetrahydrof uran 1.9 4.1 6.2 10.6 9.1 21.7 18.2 Effect of Temperature Previous work5 has suggested a strong interrelationship between the effects of organic modifier and temperature in ion-pair R P - H P L S C . Thus, an increase in temperature at a fixed acetonitrile concentration caused5 a decrease in T values (cf., previous sections). Also, found enthalpy - entropy linear compensation effects could be rationalised by equation (12) : rT = -0.068A(AH0) + 0.04 (12) where T refers to the harmonic mean of experimental temperatures and A(AH*) is the contri- bution that substituent groups have towards the enthalpies of solute transfer at this tempera- ture T .By combining values from this previous study5 with data from our present investi- gation, we can obtain AD = 0.28A(AHo) + 0.04 (13)Anal. Proc. This expression, which explains 95.5% of the variance in the two sets of values, can be further appreciated by examination of Fig. 2. 532 ADVANCES I N APPLIED HPLC I A B C D 0 30 60 Concentration, % V N Fig. 1. Relationship between slope co- efficient, H , from equation 11 with organic modifier percentage composition. A, Tetra- hydrofuran : B, propan-2-01; C, acetonitrile ; and D, methanol. In eauation (12) the sloDe coefficient value of 1 Y UJ -J 0 0.30 0.32 0.34 T-lIK x lo2 Fig. 2. Inter-relationships between the effect of temperature (A, B) a t two different acetonitrile volume fraction concentrations (A, 0.20; B, 0.25), and the effect of acetonitrile volume frac- tion phase concentration (C) at a fixed temperature (30 "C) on the capacity ratio of benzoic acid, using alkyl- benzyldimethylammonium chloride as pairing ion. -0.068 obtained for ion-pair systems5 is consistint with a Galue (-6.076) obtained previously for non-ion-pair RP-HPLSC systems,15 suggesting that equation (12) may be general for all reversed-phase chromatography. To test this hypothesis the coefficients for the general form of equation (12) have been obtained for other types of systems using some benzoic and phthalic acids as model compounds. The results are presented in Table V. The metal chelate system showed noticeable selectivity differences compared with the other systems, particularly for the ph thalic acids,15 and also showed a greater dependence on temperature compared with the alkylbenzyldimethyl- ammonium chloride system. These results show that equation (12) is generally applicable to the reversed-phase systems examined, although much more data on the relationships between group selectivity and temperature need to be obtained before such an equation can be regarded as general, TABLE V ENTHALPY - ENTROPY COMPENSATION RELATIONSHIPS ACCORDING TO EQUATION (12) Solutes Alkyl benzoates . . . . Phase system Methanol - water/ modified silicas Slope coefficient Y Reference -0.076 0.931 15 Benzoic acids . . . . . . Acetonitrile - water/ODS - 0.068 0.976 5 alkylbenz yldimethylammonium Benzoic and phthalic acids . . Acetonitrile - water/SAS -0.075 0.920 This study H ypersil alkylbenzyldimethylammonium Benzoic and phthalic acids . . Acetonitrile - water/SAS - 0.0G8 0.833 This study H ypersil 12-dien-Zn( I I)December, 1980 RESEARCH AND DEVELOPMENT TOPICS 533 We are grateful to the Science Research Council and to Imperial Chemical Industries for providing a CASE studentship for C.M.R. 1. 2. 3. 4. 5. 6. 7 . 8. 9. 10. 11. 12. 13. 14. 15. References Tomlinson, E. Riley, C. M., and Jefferies, T. hl.. J . Chromatogr., 1978, 159, 315. Tomlinson, E., J . Chromatogr., 1975, 113, 1. Tomlinson, E., Poppe, H., and Kraak, J . C., J . Pharm. Pharmacol., 1976, 28, 43P. Tomlinson, E., Proc. Anal. Div. Chew. SOC., 1977, 14, 294. Riley, C. M., Tomlinson, E., and Jefferies, T. M., J . Chromatogr., 1979, 185, 197. Tomlinson, E., Poppe, H., and Kraak, J . C., Methodol. Surv. Biochem., 1978, 7, 207. Wells, P. R., “Linear Free-energy Relationships,” Academic Press, London, 1968. Schoenmakers, P. J., Billiet, H. A. H., and de Galan, L., J . Chromatogr., 1979, 185, 179. Hartwick, R. A., Gill, C. M., and Brown, P. R., Anal. Chem., 1979, 51, 34. Tomlinson, E., Riley, C. M., and Jefferies, T. M., J . Chromatogr., 1979, 173, 89. Iwasa, J., Fujita, T., and Hansch, C., J . Med. Chem., 1965, 8, 150. Hammett, I,. P., “Physical Organic Chemistry,” McGraw-Hill, New York, 1940. Nice, E. C., and O’Hare, M. J . , J . Chromatogr., 1979, 162, 401. Karger, H. L., Gant, T. G., Hartkopf, A., and Weiner, P. H., J . Chvomatogr., 1976, 123, 63. Tomlinson, E., Poppe, H., and Kraak, J. C . , to be published.

ISSN:0144-557X    

DOI:10.1039/AP9801700508

出版商:RSC    

年代:1980

数据来源: RSC

摘要:

December, 1980 RESEARCH AND DEVELOPMENT TOPICS 533 Research and Development Topics in Analytical Chemistry The following are summaries of five of the papers presented at the Research and Development Topics in Analytical Chemistry Meeting of the Analytical Division held on April 1st and 2nd, 1980, at the University of Kent at Canterbury. Summaries of seven other papers will be published in the January, 1981, issue of AnalyticaZ Proceedings. Development of Ion-selective Electrodes and Flow Injection Analysis for Sulphides and Thiols E. Jane Duffield, G. J. Moody and J. D. R. Thomas Chemistry Department, U WIST, Cardin, CF1 3N U The capability of ion-selective electrodes based on silver sulphide sensing membranes for monitoring low levels of sulphide-ion activity has been demonstrated by potentiometric titrations,l-5 alkaline pulp liquor monitoring the growth of DesuZphovibrio desuZphuricanss and stability constant determinati~n.~ The electrodes also respond to simple t h i o l ~ .~ , l ~ This study deals with sulphide ion-selective electrode calibration for direct measurement of sulphides and alkanethiols and use of the electrode for flow injection analysis. Electrode Response to Sulphide Ions Sulphide is susceptible to appreciable hydrolysis in the middle pH range : Sc + H20 + HS- + OH- . . . . .. * . (1) HS- + H20 + H2S + OH- . . . . . . * * (2) Only the first step affects the sulphide-ion concentration in moderately alkaline solutions. Thus, in the pH range 11.4-11.8 the sulphide ion activity, [S2-1, sensed by a silver sulphide membrane electrode, is lower than the total level, [S2-],, according to the relationship3~l1534 RESEARCH AND DEVELOPMENT TOPICS Anal.Proc. where K , is the ionisation constant of HS- and f a2- is the activity coefficient of the sulphide ion. Rather than calculate [S2-IT from this relationship, it is simpler, when interest is in total sulphide, to take direct measurements in very alkaline solutions such as 1 M sodium hydroxide solution. An antioxidant reduces oxidation problems12 and calibration in a background of 25% V/V sulphide antioxidant buffer (SAOB) consisting of sodium hydroxide (80 g dm-3), disodium ethylenediaminetetraacetate (85 g dm-3) and ascorbic acid (36 g dm-3) by adding known increments of standardised sulphide solution gives a linear response down to about 2 x lW7 M sulphide ions.Electrode Response to Thiols The slope of the response of a silver sulphide membrane ion-selective electrode to various thiols in sodium hydroxide solution was found to be about -110 mV decade-1 for cysteine, thioglycollic acid and mercaptopropionic acid and - 84 mV decade-l for mercaptoethanoLl0 These values suggest { Ag [SCH,CH( NH,) CO,] ,I3-, [Ag(SCH,CO,) ,I3-, [Ag(SCH2CH,C02) ,I3- and [Ag,(SCH,CH,OH) 3]- as possible formulae for the respective complexes.1° The nature of the mercaptopropanol complex is not obvious from the normal rules of co-ordination chemistry, but other species may be involved in complexation, such as the hydroxide ions of the background 5 M sodium hydroxide employed. Simple alkanethiols can be expected to be detected similarly by silver sulphide membrane ion-selective electrodes.However, the extreme volatility of the lower members renders calibrations difficult because of rapid evaporation when dispensing standards ; ethanethiol and propanethiol have vapour pressures of about 400 and 100 mmHg, respectively, at 25 "C. Nevertheless, the less volatile heptanethiol can be dispensed with an Oxford autopipette, and the material is sufficiently soluble in aqueous systems for it to be used as a model for more volatile counterparts. Thus, calibration of an Orion 94-16A sulphide ion-selective electrode for heptanethiol in 25% V/V SAOB under nitrogen is linear from 9 x to lo-, M with a slope of -90 mV decade-l. The upper limit is restricted by the poor solubility of the heptanethiol, but to counter this, calibration in ethanolic sodium hydroxide (20 dm-3) is linear up to 1 0 - l ~ heptanethiol with a slope of -96mV decade-l.Such slopes suggest [Ag,(SC,H,,) 3]- as the formula of the complex, similar to that of mercaptoethanol noted above. Slopes for calibrations of heptanethiol in media not containing sodium hydroxide are greater, being -125 mV decade-1 for sodium acetate (0.1 M) in a mixture of methanol (474), propan- 2-01 (92%) and toluene (4y0), -119 mV decade-l in a mixture of 2,2,4-trimethylpentane and propan-1-01 (1 + 1) and -106 mV decade-1 in ethanol. However, these slopes, rather than being attributable to a complex of different stoicheiometry, may have their origin in solvo- lysis reactions, such as in the relations discussed above for sulphide ions in water and to which the large calibration slope of the sulphide ion-selective electrode for sulphide ions in water has beer, ascribed.12 Flow Injection Analysis The sulphide ion-selective electrode may be conveniently employed for flow injection analysis, whereby the calibrating standards and samples are allowed to flow over the mem- brane surface of the electrode coupled to a reference electrode in the manner previously described for other ion-selective electrodes.13J4 Normally, the carrier solution is pumped through continuously and the injected standards or samples pass through a mixing coil before flowing over the ion-selective electrode membrane surface.For the analyses carried out here, the samples containing sulphide were already in 1 M sodium hydroxide solution and such a carrier is suitable for calibrating standards.Alterna- tively, such samples require minimal mixing with carrier when SAOB is employed so that the flow injection system can be of the simplest possible minimal dispersion design. How- ever, an unexpected problem arising during calibration with sulphide standards in 25% SAOB gave a drifting base line and irregularities in response peaks. This was attributed to the large change in e.m.f. that the large-range sulphide ion-selective electrode has to counter between standards and carrier. The effect may be countered by having a background of 1 O V 6 ~ sodium sulphide in the carrier 25% SAOB. With this modification in the carrier,December, 1980 RESEARCH AND DEVELOPMENT TOPICS 535 reproducible flow injection e.m.f.response peaks were obtained for sulphide standards in the range 104-10-1 M, which covered the 10-3-10-2 M levels of sulphide in the various samples examined. Conclusion The practical difficulties of the response of siiver sulphide membrane ion-selective electrodes to sulphides and thiols relate to oxidation and volatility, but these can largely be overcome by attention to detail in the choice of calibrating standards and in dispensing. The flow injection analysis system can give greater control in these respects and allows an important role for flow injection analysis in the control of sulphides and thiols in industrial processes; this will be a continuing objective of this study. The authors thank the Science Research Council for a studentship (to E.J.D.) under the Co-operative Awards in Science and Engineering Scheme in conjunction with Esso Petroleum Company Ltd.1. 2. 3. 4. 5 . 6. 7. 8. 9. 10. 11. 12. 13. 14. References “Instruction Manual, Sulphide Ion Electrode, Silver Ion Electrode, Model 94-16,” Orion Research Pungor, E., Anal. Chem., 1967, 39, 28A. Hseu, T. M., and Rechnitz, G. A., Anal. Chem., 1968, 40, 1054. Gruen, L. C., and Harrap, B. S., J . SOC. Leather Trades Chem., 1971, 55, 131. Naumann, R., and Weber, Ch., 2. Anal. Chem., 1971, 253, 111. Swartz, J. L., and Light, T. S., J. Tech. Assoc. Pulp Paper Ind., 1970, 53, 90. Frant, M. S., and Ross, J. W., J . Tech. Assoc. Pulp Paper Ind., 1970, 53, 1753. Crombie, D. J., Moody, G. J., and Thomas, J .D. R., Lab. Pract., 1980, 29, 259. Gruen, L. C., and Harrap, B. S., Anal. Biochem., 1971, 42, 377. Tseng, P. K. C., andGutknecht, W. F., Anal. Chem., 1975, 47, 2316. Vesself, J., Jensen, 0. J., and Nicolaisen, B., Anal. Chim. A d a , 1972, 62, 1. Crombie, D. J., Moody, G. J . , and Thomas, J. D. R., Anal. Chim. A d a , 1975, 80, 1. RbiiCka, J., and Hansen, E. H., Anal. Chim. Acta, 1975, 78, 145. Hansen, E. H., Ghose, A. K., and RbiiCka, J., Analyst, 1977, 102, 705. Inc., Cambridge, Mass., 1970. Recent Developments in Flow Injection Analysis : Determination of Bismuth, Thorium and Copper with Pyrocatechol Violet by Exploitation of pH S. Baban Department of Chemistry, University College of Swansea, Singleton Park, Swansea, S A 2 8PP Flow injection analysis (FIA) has been developed as a simple, accurate and rapid method of analysis. The method is based on the rapid injection of an aqueous sample into a flowing carrier stream of reagent without air segmenta- tion.In this paper, the usefulness of FIA for the determination of bismuth, thorium and copper by the formation of complexes with pyrocatechol violet (PCV), which are pH dependent, is demonstrated. It is well known that PCV forms coloured compounds with many metals, the most stable complexes formed being for bismuth and thorium in the pH range 2-6, and for copper at about pH 7 in the presence of pyridinea2 Because of the absence of air segmentation in FIA it is possible to create pH gradients over the sample plug as it mixes with the ~ a r r i e r . ~ Therefore there is the possibility that pH - absorbance curves may be obtained under the dynamic conditions of FIA and rapid multi-element analyses may be performed.Typical sampling rates are 120-150 per h0ur.l Experimental Reagents Carrier soZutioas. A 5 x M solution of pyrocatechol violet was prepared. For buffers of pH below 2 the pH was controlled by addition of nitric acid, for pH 2-5 standard phthalate buffer was used, and for pH 6-7 pyridine - sodium hydroxide was used.4536 RESEARCH AND DEVELOPMENT TOPICS Anal. Proc. M stock solution of bismuth, thorium and copper was prepared, standardised gravimetrically, and diluted to give solutions of the metals with concentrations in the range 3 x 10-5-10-4~. Standard samples. A Apparatus Fig. 1 is a schematic diagram of the apparatus used.A Desaga 132100 multi-purpose peristaltic pump was used, which provided a constant flow-rate. The samples were injected from a disposable plastic syringe (1 ml) through the septum valve into the reagent stream. Polyethylene tubing of 0.86mm i.d. was used for the various coils of the system. The reaction coil and the mixing coil were 50 and 100 cm long, respectively. The detector used was a phototransducer using a yellow LED (maximum emissivity 590 nm) as the light source, and a silicon phototransistor as the sensing element. The light components were glued into a prepared Perspex block, such that the light path of 1.5 mm was normal to the direction of the f l o ~ . ~ ~ ~ The current signal from the phototransducer was fed to an amplifier circuit, which produced an output voltage proportional to the intensity of light falling on the photo- transistor.This signal was fed to a chart recorder. ml min-’ Sample (0.2 ml) L / / r 50 cm //I 100 cm Ry+ Buffer Pump Detector pH meter Waste 3 elu Amplifier Recorder Fig. 1. Schematic diagram of flow injection analysis system for the determination of bismuth, thorium and copper with pyrocatechol violet. All coils are of 0.86 mm i.d. Procedure solution were injected. The carrier flow-rate was adjusted to 2 ml min-l, and aliquots of 200 pl of the sample The resultant coloured complexes formed were measured. Results and Discussion Effect of pH It is well known that the formation of complexes of metal ions with a spectrometric reagent, especially those ligands which may protonate, e.g., PCV, is dependent on the pH of the reaction medium and the nature of the solvent and ligand concerned.Experiments with the three metals and the pH - absorbance curves obtained show that PCV forms coloured compounds with bismuth and thorium. The absorbance maxima occur in the pH range 34, as do the most stable complexes. However, with copper in the presence of pyridine the absorbance maximum occurred in the pH range 6-7, showing that a complex had formed. In both instances, the formation of a complex was seen by a sharp colour change from yellow to blue. Sensitivity and Precision Calibration graphs were obtained for bismuth, thorium and copper under the pH con- ditions previously mentioned. An example of a calibration graph for bismuth is shown in Fig.2. Successive sample injections of progressively increasing concentration followed by progressive decreases in the concentration resulted in the symmetrical pattern of peaks shown in Fig. 2. Reproducible peak heights were obtained for samples of the same concentration. The sensitivity for each was 3 x 10-5-10-4 M.December, 1980 RESEARCH AND DEVELOPMENT TOPICS 537 g::V .- QJ f3 p m r Y 80 w 0) 2 f L 0 2 4 6 8 1 0 Concentration XI O ~ / M J 8 8 7 1 I 7 I 4 4- Time Fig. 2 . (a) Calibration graph for bismuth determination; (b) peaks resulting from injection of successive samples whose concentration steadily increased then decreased. Conclusions This work demonstrates the way in which several different elements can be determined using the same reagent, merely by changing the pH of the reaction medium.The results show that the pH is a critical factor governing the formation of complexes. The effect will be exploited in the form of a pH gradient in the flowing carrier to permit multi-element determinations on a single sample. This method may also be applied to the rapid analysis of metals from different source materials. The author is grateful to the University of Sulaimanya for a grant for carrying out this research. References 1. RhiiEka, J., and Hansen, E., Anal. Chim. Acta, 1975, 78, 145. 2. Vogel, A. I., “Textbook of Quantitative Inorganic Analysis,” Fourth Edition, Longmans, London, 3. Betteridge, D., and Fields, B., Anal. Chim. Acta, submitted for publication. 4. Weast, R. C., Editor, “Handbook of Chemistry and Physics,” Fifty-third Edition, CRC Press, 5 .1978. Cleveland, Ohio, 1972-73. Betteridge, D., Dagless, E. L., Fields, B., and Graves, N. F., Analyst, 1978, 103, 897. Application of High-performance Liquid Chromatography and Enzymatic Methods to Food Analysis with Special Reference to the Determination of Sugars P. A. Jackson Laboratory of the Government Chemist, Cornwall House, Stamford Street, London, SE1 9NQ The increasing complexity of processed foods has resulted in a need for highly specific methods. of analysis. High-performance liquid chromatography (HPLC) and enzymatic techniques have proved to be particularly useful in this respect and have been applied to a wide variety of foodstuffs. This paper is specifically concerned with the application of HPLC and enzymatic methods to the determination of the major sugars found in processed foods.A large number of techniques for the determination of sugar are currently in use, including the classical wet-chemical methods such as Fehling’s titrations, optical methods including538 RESEARCH AND DEVELOPMENT TOPICS Anal. Proc. polarimetry and refractometry, and various chromatographic techniques, such as gas - liquid chromatography, thin-layer chromatography, paper chromatography and conventional liquid chromatography. HPLC and enzymatic methods offer significant advantages over all of these techniques in terms of short analysis times and high specificity and, with enzymatic techniques, higher sensitivity. High-performance Liquid Chromatography In the last 5 years many workers have reported the use of chemically bonded stationary phases in the HPLC separation of carbohydrates, One of the first major advances in the use of these columns occurred in 1974 when Waters Associates (Hartford, Cheshire) marketed a carbohydrate column.Many workers have subsequently used this ~olurnnl-~ but even at optimum operating conditions this laboratory was unable to achieve complete separation of glucose and fructose. In addition, it was observed that the efficiency of separation of the column deteriorated rapidly ; this was attributed to the complex nature of the samples injected into the system. In 1976 the preparation of a stationary phase consisting of silica modified by 3-amino- prop yltrie t hoxysiloxane was reported by Schwarzenbach .4 Columns containing this station- ary phase were prepared and a significant improvement in the efficiency of separation of the sugars was obtained. This column deteriorated as quickly as the Water Associates column but it was a relatively simple procedure to prepare and pack new columns.More recently, the technique of coating silica columns with amine groups in situ, as described by Ait~etmuller,~ has been investigated. In this technique a polyfunctional amine is dissolved in the mobile phase and this continually regenerates the stationary phase as it is pumped through the system. These columns achieved a high efficiency of separation, as shown in Fig. 1, and were found to display greater long-term stability compared with the earlier columns. Calibration graphs for fructose, glucose, sucrose, lactose and maltose were found to be linear in the range 0.5-100g1-1 and repeated injections of a standard solution. of glucose gave a co- efficient of variation of approximately 2%.A CI .- - i D 1 E F Fig. 1. Illustration of efficiency achieved with silica columns coated in situ with Peaks: A, water; B, fructose: C, glucose; D, sucrose; E, maltose: and F, amine groups. lactose. Number of theoretical plates = 16435 per metre. This technique is now being used regularly in this laboratory and has been applied success- fully to a variety of analytical problems, such as the detection of small amounts of invert sugar in sucrose and the identification and determination of glucose syrups in processed foods. Although HPLC has solved many problems in the separation and determination of specific sugars in composite foods, the technique is insufficiently sensitive for certain applications.Enzymatic techniques offer increased sensitivity compared with HPLC and have been used to solve some of these problems, Enzymatic Methods The major sugars that are determined enzymatically in this laboratory are glucose, fructose,December, 1980 RESEARCH AND DEVELOPMENT TOPICS 539 sucrose and lactose. These are determined using the methods developed by Bergmeyer,G which utilise the nicotinamide adenine dinucleotide phosphate (NADP) dependent dehydro- genase enzymes. The reactions are performed entirely in spectrophotometric cuvettes and are followed by monitoring the appearance of the reduced form of nicotonamide dinucleotide phosphate (NADPH) by the absorption at 340 nm.Glucose and fructose are phosphorylated to the corresponding hexose-6-phosphate by adenosine-5’-triphosphate (ATP) and hexokinase ; the glucose-6-phosphate formed is then oxidised with glucose-6-phosphate dehydrogenase and NADP to give the gluconate-6-phos- phate and NADPH. The formation of NADPH causes an increase in absorption at 340 nm, which is proportional to the amount of glucose present in the sample. When these reactions are complete the fructose-6-phosphate in the cuvette is converted into glucose-6-phosphate by the addition of phosphoglucose isomerase , the glucose-6-phosphate formed being immediately oxidised by the enzymes already present in solution to give a second rise in absorption, which is proportional to the amount of fructose present in the original sample.Lactose may be determined in the same sample by addition, of P-galactosidase to the cuvette after the glucose and fructose determinations are complete. This enzyme specifically hydro- lyses lactose to glucose and galactose; the glucose formed immediately undergoes the same reaction as the glucose that was originally measured. This results in a third rise in absorption, which is proportional to the amount of lactose in the sample. Sucrose is determined by carry- ing out a parallel series of reactions to those for glucose after the sucrose in the sample has been hydrolysed by the addition of invertase. The amount of glucose formed on hydrolysis is directly proportional to the amount of sucrose originally present.These methods of determination are linear in the range 5-100 mg 1-I and precisions of 1-2y0 are regularly obtained. The major disadvantages of enzymatic methods are that it is im- possible to perform quantitative scans of a mixture of sugars and it is preferable to know which sugars are present before enzymatic determinations can be performed. Another disadvantage of this technique is the relatively high cost of the determination when using purified enzymes. This cost is being reduced by the use of immobilised enzymes in different types of automated sugar analysis equipment. These may be either the conventional, continuous flow AutoAnalyzer type of instrument, in which the enzyme may be immobilised on the walls of reaction coils, or the new, discrete type of instrument in which the immobilised enzymes act as selective agents in electrochemical sensors, e.g., the YSI sugar analyser (Yellow Springs Instrument Co.Inc., Yellow Springs, Ohio). Automation of sugar determinations using an AutoAnalyzer system was unsuitable for use at this laboratory, because of the wide variety of samples that are analysed, but a discrete type of analyser (YSI) has recently been purchased. The major application of this sugar analyser in the laboratory has been in the determination of lactose in milk products. A series of skimmed milk powders was analysed using this instrument, the manual enzymatic methods and the traditional Fehling’s titration; the results obtained are shown in Table I. There was TABLE I COMPARISON OF RESULTS FOR ANALYSIS OF MILK POWDERS Results are as a percentage of lactose.Sample No. 1 2 3 4 5 6 7 8 9 10 Mean . . .. Standard deviation . . Standard error of mean . . Manual enzymatic method 46.1 47.5 48.4 50.7 51.2 49.4 48.7 49.5 49.7 49.1 49.0 1.5 0.47 Immobilised enzymatic methbd 44.9 48.9 50.5 49.8 51.1 50.0 48.3 50.8 49.4 49.4 49.3 1.8 0.56 Fehling’s titration 51.7 50.8 52.3 49.5 51.9 50.4 47.0 48.1 48.7 49.5 50.0 1.7 0.55Anal. Proc. 540 RESEARCH AND DEVELOPMENT TOPICS no significant difference between the results obtained by the two enzymatic techniques, but it was found that the Fehling’s titration tended to give slightly higher results than enzymatic analysis. Conclusion In summary it is considered that HPLC and enzymatic methods are complementary tech- niques.The main advantages of HPLC are fast analysis time and selectivity. The more common sugars are completely separated and individually determined in 10 min. The major disadvantage of the technique is the lack of sensitivity, the limit of sensitivity being 0.5 g 1-l. In comparison, enzymatic analysis is more sensitive, the lower limit of sensitivity being 5 mg 1-l. A variety of samples have been analysed using both techniques and results are in good agree- ment, as shown in Table 11. TABLE I1 COMPARISON OF HPLC AND ENZYMATIC METHODS Results are as a percentage of sugar in the sample. Enzymatic Sample Sugar method HPLC Boiled sweet . . . . . . Redcurrant jelly . . .. . . Biscuits . . . . . . .. Toffee . . . . . . . . Cheese powder . . . . . . Raisins .. . . .. . . Dates . . . . .. . . Sultanas . . .. . . . . Glucose Sucrose Fructose Glucose Sucrose Sucrose Fructose Glucose Sucrose Lactose Lactose Fructose Glucose Fructose Fructose Glucose 12.0 50.5 16.5 16.6 33.7 44.8 5.3 6.6 40.1 2.6 , 24.5 38.5 36.3 34.2 36.3 36.3 11.2 49.1 16.3 17.3 32.9 44.7 5.1 6.7 41.4 2.5 25.1 39.2 36.4 35.8 38.0 36.0 If a sample with an unknown sugar content is to be analysed it is initially injected on to the HPLC system where the component sugars are identified and determined. If a more accurate analysis is then necessary the sample is analysed by an enzymatic method for each specific sugar. References 1. 2. 3. 4. 5. 6. Palmer, J. K., Anal. Lett., 1975, 8, 215. Linden, J. C., and Lawhead, J., J . Chromatogr., 1975, 105, 125. Hurst, W.J., and Martin, R. A., J . Assoc. 08. Anal. Chem., 1977, 60, 1180. Schwarzenbach, W., J . Chromatogr., 1976, 117, 206. Aitzetmuller, K., J . Chromatogr., 1978, 156, 354. Bergmeyer, H. U., “Methods of Enzymatic Analysis,” Second Edition, Academic Press, New York, London, 1974. Trace Determination of the Faecal Sterol Coprostanol David V. McCalley, Michael Cooke and Graham Nickless Department of Inorganic Chemistry, The University, Bristol, BS8 1 TS The assessment of sewage pollution in the environment is of prime importance for health, aesthetic and ecological reasons. As pathogenic bacteria and viruses are very difficult to isolate and enumerate, faecal pollution is generally assessed via an indicator substance, either microbiological or chemical, which is present in sewage in larger amounts.December, 1980 RESEARCH AND DEVELOPMENT TOPICS 541 The method in general use at present is microbiological, involving the enumeration of faecal coliform organisms which are present in large numbers in sewage.However, this method presents some problems in that faecal coliforms may be destroyed under conditions of tempera- ture, sunlight and salinity in which pathogens may survive. Considerable scope therefore exists for the development of a chemical indicator of pollution with a longer survival period in the environment. Coprostanol (5P-cholestan-3/3-01), which is unique to the faeces of man and higher animals, has recently been proposed as a suitable indicator. The analytical problem is therefore to determine coprostanol at trace levels in the environment, involving its separation from numerous naturally occurring steroids, especially cholesterol, which is not unique to sew age.Firstly, the separation of standards, especially of coprostanol and cholesterol, was attempted by packed and capillary column gas - liquid chromatography (GLC) and by high-performance liquid chromatography (HPLC). Secondly, these results were combined with further investi- gations into extraction and clean-up procedures for the determination of steroids in environ- mental samples. Experimental Gas - Liquid Chromatography For flame-ionisation detection (FID) the trimethylsilyl (TMS) ethers of the sterols were pre- pared by reaction with bistrimethylsilylacetamide - trimethylchlorosilane (5 + 1) (3 h, room temperature).For electron-capture detection (ECD) the heptafluorobutyrate esters of the sterols were prepared by reaction with heptafluorobutyric anhydride using trimethylamine as catalyst. Packed columns (1.5 m x 0.4 mm i.d.) were filled with 3% OV-17 coated on Gas- Chrom Q. Capillary columns (20 m x 0.3 nim i.d.) were drawn from Pyrex glass tubes. The columns were leached with hydrochloric acid and deactivated by silylation prior to coating with SE-30 Ultraphase using the static method. Analyses were performed with splitless injection, using a temperature programme from 100 to 250 "C at 6 "C min-I. The columns were operated isothermally at 250 "C. High-performance Liquid Chromatography Benzoate and substituted benzoate derivatives were prepared by reaction of sterols with benzoyl chloride, p-nitrobenzoyl chloride or 3,5-dinitrobenzoyl chloride in pyridine (60 "C, 1 h) .Analysis was attempted on microparticulate silica columns (20 cm, 5 pm irregular particles) or octadecylsilyl bonded phase columns (10 cm, 5 pm spherical particles). Extraction and Clean-up of Samples Sediments and sewage sludges were freeze-dried and Soxhlet extracted for 24 h with chloroform - methanol (2 + 1). Liquid samples were extracted with methylene chloride. Heavily contaminated liquid effluents were first filtered through glass-fibre filter-papers (GF/C) and the particulate matter was analysed separately. Crude extracts were purified by thin-layer chromatography (TLC) on silica gel plates (0.4 mm thickness) using chloroform as the mobile phase.Results Preparation of sterol TMS ethers for gas chromatography improves the separation and eliminates losses due to column adsorption via the free hydroxyl group. Packed column GLC gave base-line separations of coprostanol, cholesterol and the internal standard 5cc- cholestane. However, under isothermal conditions, the peaks eluted close to the solvent front. Temperature-programmed analyses were not possible owing to base-line instability caused by bleeding at the high temperature and high amplifier settings necessary for the trace determina- tion of sterols. The detection limit of this method was about 5 ng of injected coprostanol. These problems were overcome by the use of capillary columns, which gave much improved separations. The coprostanol and epicoprostanol peaks were separated down to the base line on this column (Fig.1)-these compounds differ only in the orientation of the hydroxyl group to the ring. SE-30 Ultraphase columns were stable up to 350 "C with very low bleed levels and gave detection limits in the picogram range. The use of a microcell ECD with this column gave542 RESEARCH AND DEVELOPMENT TOPICS Anal. Proc. similar separations of sterol heptafluorobutyrate derivatives with even lower detection limits. However, it was later found that analyses of environmental samples suffered from interferences due to the response of the ECD to other components in the samples. Therefore, preparation of TMS derivatives and detection by FID was the preferred method. Based on the internal standard, 5ct-cholestane, the response of the FID was linear for 0-25 ng of injected coprostanol.As coprostanol has no natural chromophores, the strongly absorbing benzoate and substituted benzoate derivatives were prepared. Sterol $-nitrobenzoates were preferred owing to their absorption at 254 nm. However, both benzoate and 3,5-dinitrobenzoates were readily synthesised, and their identities were confirmed by mass spectrometry, despite accounts of difficulty experienced in the preparation of cholesterol-3,5-dinitrobenzoate reported e1sewhere.l It was found that these derivatives were fairly non-polar, and in normal-phase chromatography on silica columns rapid elution of sterols occurred with no separation, even with solvents such as hexane. Reversed-phase HPLC yielded improved results, and good peak shapes, with detection limits similar to those of packed column GLC, were obtained using pure methanol as solvent (Fig.2). However, this technique failed to produce good resolution of sterols, even failing to resolve coprostanol and cholesterol completely. Attempts to improve the separation by adding water to the mobile phase were unsuccessful as the peaks collapsed to broad humps even when only small amounts of water were added. The use of other solvents such as acetonitrile gave no improvement in separation. For analysis by HPLC, ultraviolet detection was chosen for its sensitivity. C B t I 100 250 Temperature/"C Fig. 1. Separation of a sterol mixture (as TMS derivatives) by capillary column GLC. Column : 20 m SE-30 Ultraphase. Carrier gas : hydrogen, flow-rate 2 ml min-l.Temperature programme : from 100 to 250 "C a t 6 "C min-l. Peaks: A, 5a- cholestane (internal standard) ; €3, coprostanol; C , epicoprostanol; D, cholesterol ; E, campesterol ; and F. ,!?-sitosterol. 3 3 I 0 10 Ti me/m in Fig. 2 . Analysis of sterols (as p-nitrobenzoate derivatives) by reversed- phase HPLC. Column: 10 cm Hypersil ODS. Solvent : methanol. Flow-rate: 1 ml min-l. Peaks : A, coprostanol ; and B, cholesterol. The use of preparative TLC as a clean-up procedure permitted the removal of the correspond- Also, ing keto-steroids, which can interfere even when using high-resolution capillary columns. much of the polar material that can damage columns is eliminated by this procedure. Discussion Analysis of sterol ThlS derivatives on capillary columns using an FID was established as the method of choice for the analysis of environmental samples for sterols.Using this method together with the described extraction and clean-up procedures, good recoveries (about 90%)December, 1980 RESEARCH AND DEVELOPMENT TOPICS 543 were obtained from the analysis of spiked sediment samples. The coefficient of variation obtained from the analysis of a series of ten sediment subsamples was only 4%. Analysis of samples by combined GLC and mass spectrometry confirmed the identity and purity of the coprostanol peak in a wide variety of sample types, and also permitted the identification of many of the co-extracted components. The principal sterols in crude sewage sludge were found to be coprostanol, cholesterol, methylcoprostanol, ethylcoprostanol and other alcohols, although marine sediments were considerably more complex.Treatment of sewage sludge by digestion was found to produce epicoprostanol, although the exact nature of this reaction was unclear. Levels of coprostanol found in Severn Estuary sediments are given in Table I and indicate considerable pollution by sewage in the upper reaches. However, whereas indicator methods such as the determination of coprostanol or enumeration of faecal coliforms can give estimates of the amount of sewage pollution, the significance of these levels can only be interpreted through epidemiological studies. It should be borne in mind that although sewage can be a dangerous carrier of disease, the number of pathogens in particular samples depends very much on the general state of health of the community from which the sewage is derived.TABLE I LEVELS OF COPROSTANOL FOUND AT VARIOUS SITES IN SWANSEA BAY (SITES 1-5) AND THE SEVERN ESTUARY (SITES 6-10) Site No. Location Coprostanol, p.p.m. dry mass 1 2 3 4 5 6 7 8 9 10 Oystermouth Bay Mumbles Head West Cross Swansea University Brynmill Aust Severn Beach Portishead Weston-super-Mare Watche t (Bristol sewage sludge) 2.1 1.6 1.8 5.2 2.8 0.9 1.2 3.1 1.2 1 .o (2 500) Reference 1. Fitzpatrick, F. A., and Siggia, S., Anal. Chem., 1973, 45, 2310. Catalytic Micro-determination of Osmium at the Sub-parts per 109 Level I. N. C. Ling and G. Svehla Depavtment of Analytical Chemistry, The Queen’s University of Belfast, Belfast, BT9 5AG, N . Ireland For the determination of trace amounts of osmium we have utilised the slow reaction between hydrogen peroxide and cyanide.At pH 11.2, this reaction obeys the over-all stoicheiometry H202 + CN- --f CNO- + H20 The reaction itself is very slow (the second-order rate constant1 at room temperature is as low as 1 x 10-3 1 mol-1 s-I), but can be speeded up by adding a well defined, small amount of copper. Under such circumstances, new reaction paths are opened up, the rate-deter- mining step being a fast equilibrium followed by a reaction between a copper(1) - hydrogen peroxide complex and copper(I), bound in a number of cyanide complexes of various compositions. By kinetic measurements a third-order rate constant could be determined with the value 3.03 x lo4 l2 mok2 s-l.The over-all stoicheiometry of the process remains the same as shown above.544 RESEARCH AND DEVELOPMENT TOPICS Anal. Proc. This copper-catalysed reaction between Wydrogen peroxide and cyanide can be catalysed further by trace amounts of osmium. In the presence of the latter yet another reaction path becomes possible, which is again faster than the copper-catalysed reaction. On the basis of kinetic measurements we were able to postulate a mechanism, with a second-order rate-determinating step, characterised by a rate constant of 5.07 x lo4 1 mol-1 s-1. The kinetic study was extended to measurements at different temperatures, and so the entropies and enthalpies of activation of both the uncatalysed and catalysed reactions were determined. Two reagent solutions are made up: solution A (0.395g KCN + 0.0543g CuS0,.5H20 + 71.6g KH2P04 + 32.0g NaOH per litre) and solution B (1.176 x mol 1-1 H202). To 10 ml of a neutralised sample [containing 0-1 p.p.b. (parts per log) of osmium] are added 10ml of solution A, and potentiometric or amperometric monitoring is started. With constant stirring, 10 ml of solution B are then added and the potential (measured between platinum and saturated calomel electrodes) or the diffusion current (measured at zero potential difference between rotating platinum and saturated calomel electrodes) is recorded as a function of time. A well defined peak on either of these curves indicates the time needed for the removal of a well defined amount of cyanide. The reciprocal reaction times, l/.fL, multiplied by the blank reaction time, ti, when plotted against the analytical concentration of osmium, provides a straight line, which serves as a calibration graph for the determination (see Fig. 1). The analytical procedure is very simple. 3 1 0 0.5 1 cost P.P.b. Fig. 1. Calibration graph for determination of osmium. A detailed study of interferences was undertaken and it was found that in the determina- tion the following amounts of foreign metals can be tolerated (without causing an error greater than 5%) : 0.5 p.p.m.: Ni, Mn and (excess) Cu; 5 p.p.m.: Co, Mo, W, Al, Cr, Ru, Au, Ag, Rh; 50 p.p.m.: V, Pd, Ti, Hg, Ce, Fe(II1); 500 p.p.m.: Zn, Fe(II), U, Pt, Pb, Ba, Cd, La, Ir, Sb, Sn, Sr, Ca, Mg, Y, Zr, Se, Na, K. A detailed description of the method, together with the kinetic study of the reactions involved, will be published elsewhere.2 References 1. 2. Masson, O., J . Chem. Soc., 1907, 91, 1449. Ling, I. N. C., and Svehla, G., Talanta, in the press.

ISSN:0144-557X    

DOI:10.1039/AP9801700533

出版商:RSC    

年代:1980

数据来源: RSC

摘要:

December, 1980 EQUIPMENT NEWS 545 Equipment News Mass Spectrometer The Superspec 600, a microprocessor-controlled mass spectrometer, can analyse up to any 16 components in a gas stream, with a fast response time. Scientific Products Ltd. Insert 501 on the Reader Enquiry Service form for further information. Scanning Electron Microscope The Stereoscan 250 incorporates as standard autobrightness signal control, high-resolution record unit, split-screen imaging and a single- control “Optibeam” for routine electron optics. Also available is optional oil-free turbomole- cular pumping. A surface analysis accessory, SIM5, can be interfaced with the Stereoscan 250. This con- sists of an ion gun of 35 pm minimum beam diameter, a quadrupole mass analyser with a mass range of 1-300 and a special energy filter.Three operational modes, positive and negative SIMS and residual gas anaiyses, are automatically pre-timeable. Insert 502 on the Reader Enquiry Service form for further information. Cambridge Instrument Co. Ltd. Image Analysis System The Quantimet 800 is a fully integrated auto- matic image analysis system, which includes a detector to provide full-speed boundary-finding algorithms. All usual field-specific measure- ments can be performed. Control is effected conversationally via the user’s keyboard. Insert 503 on the Reader Enquiry Service form for further information. Cambridge Instrument Co. Ltd. Spectrofluorimeters Spectrofluorimeters are available in two models, one a single scan speed with fixed band pass, and the second with four forward and fast reverse scan speeds, with variable band pass from 1 to 32nm.Both incorporate a digital display with energy bar graph and analogue and BCD outputs available for recorders and printers. Baird-Atomic Ltd. Insert 504 on the Reader Enquiry Service form for further information. Holographic Gratings “Blazed” holographic gratings are now avail- able with increased efficiency a t a specific wavelength in addition to the usual features of standard holographic gratings. EDT Research. Insert 505 on the Reader Enquiry Service form for further information. Ultraviolet - Visible Spectrophotometer The HP 8450 A is a general purpose ultraviolet - visible spectrophotometer, utilising a reversed- optics technique by passing the light (200-800 nm) through the liquid sample to be analysed.The spectrum covering the full range is measured and displayed in 1 s. A 16-bit microcomputer with 88 000 bytes of memory controls the instrument. Hewlett-Packard Ltd. Insert 506 on the Reader Enquiry Service form for further information. Analyser The Scintrex Model UA-3 is designed for field or laboratory analysis of uranium from 0.05 x Using an electro-optical tech- nique, uranium is measured by the enhanced luminescence in the presence of a proprietary reagent. Excitation is by built-in pulsed nitrogen laser. Insert 507 on the Reader Enquiry Service form for further information. to 0.3%. Techmation Ltd. Ultraviolet - Visible Spectrophotometer The DU-8 ultraviolet - visible spectrophoto- meter comprises wavelength scanning, gel scanning, kinetic analyser, sipper and T , sys- tems.It can be programmed by plug-in cassettes. Reckman-RIIC Ltd. Insert 508 on the Reader Enquiry Service form for further information. Headspace Analyser The HS 250 is designed as an accessory to a gas chromatograph, in particular the Carlo Erba 2000 and 4000 series. It can be used with packed and WCOT or SCOT capillary columns. The specified ranges are 30-120 O C , sample sizes 2.5 ml vapour, 1-2 g solid and 0.5-5 ml liquid. Insert 509 on the Reader Enquiry Service form for further information. Erba Science (UK) Ltd. Mercury Vapour Analyser The Model 401, an economy version of the Model 301 multi-purpose mercury detector, retains the gold film sensing technique. It is sensitive to mercury vapour down to 2 p g m-3, free from interferences common to atomic- absorption techniques.546 EQUIPMENT NEWS Anal.Proc. D. A. Pitman Ltd. Insert 510 on the Reader Enquiry Service form for further information. Humidity Sensor h dew-point humidity sensor that operates a t greater than 94% R.H. is composed of a high- sensitivity resin containing dispersed carbon, which swells and increases resistance with increasing humidity. The operational range is -10 to 60 "C and O - l O O ~ o R.H. with a life of more than 500 cycles. Another humidity sensor that operates by resistance change through a hydration - dehydration process in a magnesium chromate - titanium dioxide ceramic is under development. Matsushita Electronic Components Co. Ltd. Insert 511 on the Reader Enquiry Service form for further information.Gas Sensors A sensor for liquid natural gases uses n-type semiconductor material composed chiefly of iron(II1) oxide. It is particularly sensitive to methane and isobutane gas, but relatively insensitive to smoke and alcohol vapour. Alarm concentrations range from 1000 p.p.m. for isobutane to 3 000 p.p.m. for methane and about 6 000 p.p.m. for ethanol a t 20 "C, 60% l3.H. A sensor designed for propane gas, EGS-S 130P02, is also very sensitive to isobutane, making it useful for LPG units. Matsushita Electronic Components Co. Ltd. Insert 512 on the Reader Enquiry Service form for further information. Analyser The G-300 automatic analyser can be pro- grammed for up to 30 parameters from a list of 75 tests, thus enabling the bulk of routine and emergency analyses in a clinical-chemical laboratory to be effected automatically.The list includes electrolytes, metabolites, enzymes, lipids, immunochemistry and enzyme immuno- assays. Greiner Electronics AG. Insert 513 on the Reader Enquiry Service form for further information. Concentrator The Series 320 Chemical Data System gas chromatography concentrator, which includes a microprocessor, uses the purge and trap technique to concentrate trace contaminants in water or air. It then reproducibly injects the concentrated sample into any GC, GC - MS or GC- IK system with capillary or packed columns. Kemtronix (UK) Ltd. Insert 514 on the Reader Enquiry Service form for further information. Conductivity Meter The Model 5007 is a battery-operated conduc- tivity meter calibrated in $3 cm-l to measure the electrical conductivity of liquids, slurries or pastes from 0.01 to 10000 pS cm-I.Two measuring frequencies, 70 H z and 1 kHz, are used to eliminate polarisation and capacitive effects. Insert 515 on the Reader Enquiry Service form for further information. EIL Analytical Instruments. Portable Dose-rate Meter A portable dose-rate meter, the LB133, measures radiation emission from gamma- or X-rays ranging in energy from 25 keV to 2 MeV. The radiation is detected by an energy- compensated aluminium proportional counter with a cross-section of 20 cm2. Laboratory Impex Ltd. Insert 516 on the Reader Enquiry Service form for further information. Isotope Calibrator The Pitman, Model 238, isotope calibrator has been replaced by the improved Model 270.Measurements can be made within the energy range 20 keV to 3 MeV and displayed over an indicating range of 0.001 to 100 000 MBq.December, 1980 EQUIPMENT NEWS 547 Measurement periods are 0.5, 5.0 or 50.0 s, Temperature Alarm accurately controlled by a quartz time base. The Alpha 1 Temperature Alarm can accept up to three standard AWL probes which are continually sequentially monitored. The range is -50 "C to 150 "C with an accuracy of & 0.5 "C. High- and low-temperature alarms can be set with out-of-limit conditions indicated by a red LED and an audible buzzer. D. A. Pitman Ltd. Insert 517 on the Reader Enquiry Service form for further information. Refractometer The Reichert- Jung Abbk refractometer is now available with a binary coded decimal parallel output system, which enables it to be inter- AWL Electronics Ltd- Insert 522 on the faced with data processing equipment. Reader Enquiry Service form for further Reader Enquiry Service form for further information.Resistivity Monitors Reichert- Jung UK. Insert 518 on the information. The Style B, 920 Series monitors feature dual- Digital Thermometer The range of portable digital thermometers has been extended to include new intrinsically safe (BASEEFA certified) models. Model KM2008 has a range of -30 to 800 "C with 1 "C resolution, and Model KM2002 a range of -30 to 199.5 "C with 0.5 "C resolution. Intrinsically safe probes for surface, air and liquid measurement are available. Insert 519 on the Reader Enquiry Service form for further information.Kane-May Ltd. Thermocouple Selector Module A panel-mounted thermocouple selector mod- ule, Type 7590, is designed to switch up to six pairs of thermocouples manualiy. The switch is of the break-before-make variety. Jenway Ltd. Insert 520 on the Reader Enquiry Service form for further information. Temperature Probes Temperature measurement without the neces- sity for a separate indicator is facilitated by the Series 600 Multimeter probes. Type 2 is designed for measurement of liquids, semi- solids, air or gas and plastics, and Type 4 for surf ace measurements. Digitron Instrumentation Ltd. Insert 521 on the Reader Enquiry Service form for further information. thermistor compensation to cover all ranges, a built-in manual calibration check, with a read- out meter accurate to within 2% full scale, for the measurement of resistivity of pure and ultra-pure water.Foxboro Analytical. Insert 523 on the Reader Enquiry Service form for further information. Recorder The REC 80 Servo-Graph servo-recorder is driven by a stepping-motor and has a reversible chart feed between 1 s cm-l and 2 h cm-l. The response time of the pen is less than 0.6 s. Insert 524 on the Reader Enquiry Service form for further information. Radiometer A/S. Recorder The Biomation Model 2805 master/slave wave- form recorder is a high-speed, multi-channel (up to eight) simultaneous recorder, at rates up to 5MHz with amplitude resolution of 8 bits. The master unit, an analogue - digital con- vertor with memory, performs dual-channel recording a t rates up to 5 MHz (0.2 ps per sample) and can manage up to three similar slave units, thus permitting up to eight channels of synchronous recording.Waveform informa- tion stored in the memory can be viewed on a cathode-ray tube monitor or oscilloscope. Gould Instruments Division. Insert 525 on548 ANALYTICAL CHEMISTRY TRUST FUND Anal. Proc. the Reader Enquiry Service form for further information. Automatic Titration System The automatic titrating and metering system operates by means of a microprocessor-con- trolled piston burette, available with four different volumes (5, 10, 20 and 50 ml). Manual operation is also possible. Schott-Gerate GmbH. Insert 526 on the Reader Enquiry Service form for further information. Photoacoustic Spectroscopy The Model OAS 400 spectrometer is available with an IEEE 488 computer interface, enabling i t to be connected directly to mini- and micro- computers for control and data processing prior to recording the completed spectrum.Soft- ware programmes for the Commodore PET microcomputer can be used further to extend the utility by acquiring up to four spectra. EDT Research. Insert 527 on the Reader Enquiry Service form for further information. Cuvette Rack ,4 moulded polypropylene rack to hold 12 spectrophotometer cuvettes of 10-mm light path is designed to avoid scratching quartz cuvettes by having only the corners of the cells in contact with the rack. Insert 528 on the Reader Enquiry Service form for further information. R. B. Radley & Co. Ltd. New Products The Re Aquant Reagent system, for determina- tion of water content, consists of a Re Aquant solvent and titrant.Being free from pyridine and sulphur dioxide, there is no odour problem, and it can be used with, e.g., ketones and aldehydes. A sharp, non-fleeting end-point is provided by a special buffer system. J. T. Baker Chemicals BV. Insert 529 on the Reader Enquiry Service form for further information. A stationary phase material for gel permeation chromatography on a preparative scale is now available from Jobin-Yvon through EDT. EDT Research. Insert 530 on the Reader Enquiry Service form for further information. Literature A booklet of Application Notes on Gel Perme- ation Chromatography produced by Jobin-Yvon is available from EDT. EDT Research. Insert 531 on the Reader Enquiry Service form for further information. Volume 2, No. 3, of “Peak” contains articles on the analysis of carbohydrates in food and of vitamins by HPLC. An Application Note on the use of GC -MS in drug analysis and a monograph on high-resolution gas chromato- graphy are also available. Hewlett-Packard Ltd. Insert 532 on the Reader Enquiry Service form for further information. h leaflet gives information on the System-Six system for chromatography, and also details and prices of the Kloehn range of syringes. Insert 533 on the Reader Enquiry Service form for further information. Fluid Eluting Devices. A publication outlines the use of the CIKA GC/IR interface, with an example of the analysis of a solvent system of five components ranging from 1.2% water to 21% triethylene Bio-Rad Laboratories. Insert 534 on the Reader Enquiry Service form for further information. glycol.

ISSN:0144-557X    

DOI:10.1039/AP9801700545

出版商:RSC    

年代:1980

数据来源: RSC

摘要:

548 ANALYTICAL CHEMISTRY TRUST FUND Anal. Proc. Ana lytica I Chemistry Trust Fund SAC Fellowships The Trustees invite proposals for research projects likely to make major contributions to the advancement of analytical chemistry in the UK. Projects with original ideas for research are particularly requested. Proposals will be considered from applicants with a first-class research background in non-academic establish- ments, such as industrial organisations, govern-TUCK IN UNDER FLAP A ANALYTICAL PROCEEDINGS December, 1980 READER ENQUIRY SERVICE For further information about any of the products featured in the advertise- $ ments or the Equipment News feature in this issue, please write the appro- 3 Postage paid if posted in the British Isles but overseas readers must affix a stamp.priate number in one of the boxes below. n I I l I . 1 i I (Please use BLOCK CAPITALS) NAME ................................................................................................................................................................. 0 C C U PATI 0 N ......................................................................................................................................... AD DRESS ..................................................................................................................................................... SECOND FOLD Postage will be Paid by Licensee Do not affix Postage Stamps if posted in Gt. Britain, Channel Islands or N. Ireland I I BUSINESS REPLY SERVICE Licence No. W.D. 106 Reader Enquiry Service Analytical Proceedings The Royal Society of Chemistry Burlington House Piccadilly London W1 E 6WF ENGLAND THIRD FOLD FLAP A55 0 INDUSTRIAL TRAVEL FELLOWSHIPS Anal.Proc. ment laboratories and Research Associations, as well as from those in academic institutions. Applications must be made by prospective Fellows only. The value of a Fellowship is related to the Lecturer scale for non-clinical academic staff. SAC Studentships The Trustees invite proposals from super- visors for research projects likely to make important contributions to the advancement of analytical chemistry in the UK and which are suitable for well qualified postgraduate students. Projects will be assessed particularly on the originality of the research proposed. Applications may be submitted by research supervisors who must be members of the Analytical Division of the Royal Society of Chemistry of at least 2 years' standing. Proposals for projects to start in the Autumn term of 1981 will be considered early in that year when a tentative award may be made, subject to the Trustees being satisfied by the Summer of 1981 that a student acceptable to them is available. The value of a Studentship is between L1565 and L2570 per annum mini- mum, according to circumstances, plus fees. Application Regulations for the Fellowships and the Studentships can be obtained from the Secre- tary, Analytical Division, Royal Society of Chemistry, Burlington House, London, WlV OBN. The closing date for applications is January 31st, 1981.

ISSN:0144-557X    

DOI:10.1039/AP9801700548

出版商:RSC    

年代:1980

数据来源: RSC

摘要:

55 0 INDUSTRIAL TRAVEL FELLOWSHIPS Anal. Proc. Analytical Chemistry Industrial Travel Fellowships For several years now the Analytical Chemistry Trust Fund has been awarding Studentships to enable suitably qualified members to undertake postgraduate research for higher degrees a t universities and polytechnics. It has also been prepared to award Postdoctoral Fellowships for more advanced work in these institutions. The Trustees have now decided that the time is ripe to utilise some of the income of the Trust Fund to further the development of Analytical Chemistry in an industrial context, so as to enable young analytical chemists to extend their knowledge and experience. The Trustees have decided to offer a limited number of Travel Fellowships in 1981 to enable AD members employed in industry, research associations or government-funded laboratories to attend important major Analytical Chemistry Meetings in Europe and the USA.I t is hoped that the award of this kind of Fellowship will stimulate analytical chemists under 30 years of age to broaden their experience by making con- tacts with other analytical chemists in different countries. I t should also aid their career development and encourage them to play a larger part in the affairs of the Analytical Division. The Trustees have decided not to lay down rigid rules for the award of these Fellowships but rather to work within fairly broad guidelines so that suitable candidates can be encouraged to make applications. Successful candidates will be encouraged to submit papers for presentation a t the scientific meeting they wish to attend.In addition, they will be expected to make contact with major manufacturers of analytical instru- ments and equipment in the area of the confer- ence or to pay visits to institutions or organisa- tions where analytical chemistry plays a major role. Candidates will be expected to have the full support of their employers and it would be advantageous if employers could make arrange- ments for holders of Fellowships to visit another industrial company in an allied field situated in the vicinity of the conference. On completion of the Fellowship the recipient will be required to make a detailed report to the Trustees of the use to which he or she has placed the Fellowship and the advantages that have accrued from its award. It is hoped to publish a summary of these submissions in Analytical Proceedings and also to invite holders of Fellow- ships to present verbal reports at one of the Division’s Group or Regional Meetings.The Trustees have not placed any financial limit on the amount of each award, but the sum allocated would be sufficient to cover the cost of travel and a contribution to the living expenses of the applicant during the period spent overseas. The Trustees have taken soundings from their own members employed in industry and also from the Chemical Industries Association as to the amount of support which employers would be willing to give to this scheme. They have been encouraged by the response and have noted with interest that industry is very keen on the competitive aspects for the allocation of any awards.December, 1980 PUBLICATIONS RECEIVED The Trustees have decided that the first Fellowships should be awarded in 1981 and that applications should be sought during the next few months. Each applicant would have to provide detailed proposals of the purpose to which the Fellowship would be placed and how it would enable him to broaden his outlook and develop his career. Applications should also include statements from the candidate’s em- ployers that they would be prepared to support the applicant, if successful, by making time available and by other means. 551

ISSN:0144-557X    

DOI:10.1039/AP9801700550

出版商:RSC    

年代:1980

数据来源: RSC

摘要:

PUBLICATIONS RECEIVED 551 alloy, ore and other analyses; and a review of X-ray spectrometric analysis on Mars, Earth and the moon. Elemental analysis of rocks and ores, general applications of XRF, mathematical methods in XRF and X-ray diffraction applica- tions are among the many other areas covered. December, 1980 Publications Received Modern Polarographic Methods in Analy- tical Chemistry. A. M. Bond. Monographs in Electroanalytical Chemistry and Electrochemistry. Pp. xviii + 518. Marcel Dekker. 1980. Price Swfr135. ISBN 0 8247 6849 3. This is a book for the analytical chemist who is considering using polarography and wishes to ascertain which polarographic method to choose and how to use it. With the rebirth of interest in electroanalytical methods, and in polaro- graphy in particular, a vast number of polaro- graphic methods have become available, some for the first time and others based on techniques recently revived. Each method has its subtle drawbacks and advantages, and it is difficult for the chemist to know how to make the best use of this recently elaborated methodology. This book provides an up-to-date discussion of modern polarographic methods, complete with examples, experimental details and relative merits of each method.Advances in X-Ray Analysis. Edited by John R. Rhodes, Charles S. Barrett, Donald E. Leyden, John B. Newkirk, Paul K. Predecki and Clayton 0. Ruud. Pp. xviii + 390. Plenum Press. 1980. Price $45. ISBN 0 306 40435 4. This volume constitutes the Proceedings of the 1979 Denver Conference on the Applications of X-ray Analysis, the 28th in the series.The book is concerned with the subject of field applications of X-ray fluorescence with particu- lar reference to the analysis of raw materials such as rocks, ores and coal. Major subjects covered include recent advances in the applica- tion of X-ray emission techniques to geochemi- cal, borehole and on-stream analysis ; develop- ments in the use of portable instruments for Volume 23. Introduction to Modern Liquid Chromato- graphy. Second Edition. L. R. Snyder and J. J. Kirkland. Pp. xxii + 863. John Wiley. 1979. Price fll8. ISBN 0 471 03822 9. This new edition has been completely re-written to incorporate recent advances in the field, in- cluding the nearly complete solution of the ubiquitous detector problem through the use of spectrophotometer detectors operating down to 190nm, enabling the sensitive detection of almost any compound type, and increased use of fluorescence and electrochemical detectors, plus off-line and on-line derivatisation, to avoid detection problems.The book is organised in three parts : Chapters 1-6 cover the basics of LC, Chapters 7-12 dis- cuss the six LC inetliods and their applications, and Chapters 13-19 address various specialised areas. The first two sections permit the novice to master quickly the essential information required to perform actual separations and analyses. They also provide additional material for in-depth comprehension of how HPLC is carried out, what equipment is needed, and possible applications. The last section offers a thorough treatment of a number of specialised areas which are important for optimum results with this procedure.These areas include quantitative and qualitative analysis by HPLC, preparative scale separations, gradient elution and column-switching, sample pre-treatment and reaction detectors, automated systems for high-volume testing and/or samples requiring pre-treatment, troubleshooting and sample artifacts such as band-tailing, and selecting and developing an LC method(s) for a particular application. Treatise on Analytical Chemistry. Part 11. Analytical Chemistry of Inorganic and Organic Compounds. Volume 16. Func- tional Groups. R. F. Muraca. Pp. xxii + 560. John W7iley. 1980. Price A26.95; $56.55. ISBN 0 471 05857 2. Volume 16, Functional Groups, contains Section B-2, Organic Analysis 11, Functional Groups, and consists of four chapters, each by R.F. Muraca, devoted to different groups : Azoxy Group; Nitro and Nitroso Groups; Nitrate and552 PUBLICATIONS RECEIVED Anal. Proc. Nitrite Ester Groups ; and Nitrile, Isocyanide, Cyanamide and Carbodiimide Groups. Treatise on Analytical Chemistry. Part I. Theory and Practice. Second Edition. Volume 2 : Section D (continued) : Solution Equilibria and Chemistry. Edited by I. M. Kolthoff and Philip J. Elving. Pp. xxviii + 815. John Wiley. 1979. Price A39.60. Volume 2 continues Section D (Solution Equilibria and Chemistry) and contains Chap- ters 15-23 as follows: 15, Graphic Presentation of Equilibrium Data, by E. Hogfeldt; 16, Graphic and Computational Methods in the Evaluation of Stability Constants, by E.Hogfeldt; 17, Concepts of Acids and Bases, by I. M. Kolthoff; 18, Acid - Base Equilibria, Buffers and Titrations in Water, by D. Rosen- thal and P. Zuman; 19, Acid - Base Equilibria in Non-aqueous Solutions (5 sections by various authors) ; 20, Complexation Reactions, by A. Kingbom and E. Wanninen; 21, Masking and De-masking in Analytical Chemistry, by D. D. Perrin ; 22, Mechanisms of Oxidation - Reduction Reactions, by R. G. Linct; 23, Induced Reac- tions in Chemical Analysis, by L. J . Csbyi. Fundamentals of Organic and Bio- chemistry. Miriam Malm. Pp. x + 438. Van h’ostrand. 1980. Price L11.20 (softback). ISBN 0 442 26234 6. Archaeological Chemistry. A Sourcebook on the Applications of Chemistry to Archaeology. Zvi Goffer.Chemical Analysis, Volume 55. Pp. xviii + 376. John Wiley. 1980. Price j515.80. The application of the natural sciences to archaeology is a relatively recent development. This book provides archaeologists with a guide that discusses the help they can obtain from chemistry in pursuit of their field. Methods and techniques are discussed and case histories described to illustrate possible applications. There are four main sections : Chemistry, includ- ing chapters on Analytical Chemistry, Spectros- copy and Radioactivity and Nuclear Energy ; Materials and Technologies ; Decay and Restora- tion of Archaeological Materials ; and Dating. Methods for Analysis of Musts and Wines. M. A. Amerine and C. S. Ough. Pp. x + 341. John Wiley. 1980. Price Al9. ISBN 0 471 05077 6.This is a book on up-to-date and precise methods for the chemical analysis of musts and wines. ISBN 0 471 05510 7 . ISBPU’ 0 471 05156 X. First published as the wine section of the “Encyclopedia of Chemical Technology” this expanded version surveys many new methods that use more modern instrumentation to determine the optimum procedure for the more important constituents. Most of the procedures included presume fluent command of quantitative analytical chemistry as well as considerable laboratory practice. The concern with quality-control standards and legal limits on the amount of natural and added components makes this a useful book for enologists. Atmospheric Pollution 1980 Edited by Michel M, Benarie. Studies in Environmental Science 8. Pp. xvi + 440.Elsevier. 1980. Price $73.25; Dfl150. ISBN 0 444 41889 X. This book comprises a selection of 61 of the 81 papers presented a t the 14th International Colloquium on Atmospheric Pollution held in Paris, 5-8 May, 1980. The contents include Modelling; Gaussian Plume; Airflow and Dis- persion ; Analog Modelling ; Pollutant Forma- tion, Transformation and Transport ; Computa- tions and Statistical Representations; Air Chemistry and Formation of Particulate Matter ; Aerosol Physics and Measurement Concerning the Suspended Particulate Matter ; Monitoring Networks and Survey Results; and Effects on Man and on Vegetation. British Pharmacopoeia 1980. Volumes I and 11. Pp. xxxiv + 516 (Volume I) and vi + 517-940 + Al-AZ56, (Volume 11). H.M. Stationery Office. 1980. Price k60 per set.ISBN 0 11 320688 7. Elemental Analysis of Biological Materials. Current Problems and Techniques with Special Reference to Trace Elements. International Atomic Energy Agency. Techni- cal Reports Series No. 197. Pp. viii + 371. International Atomic Energy Agency. 1980. Price Sch. 530. This book is the outcome of a principal recom- mendation by an IAEA Advisory Group to produce a report containing a comparative review of selected techniques for the assay of trace and minor elements in biological materials. The sections include The Need for Trace Element Analyses in the Life Sciences, Sampling and Sample Preparation for Trace Element Analysis, Analytical Techniques for Trace and Minor Elements in Biological Materials (the chapters ISBN 92 0 115080 6.December, 1980 PUBLICATIONS RECEIVED 553 in this section were presented as Review Papers a t an IAEA Symposium on Nuclear Activation Techniques in the Life Sciences, held in Vienna in 1978) and Analytical Quality Control. Gas Chromatography with Glass Capillary Columns.Second Edition. Walter Jennings. Fp. xiv + 320. Academic Press. 1980. Price $25. ISBN 0 12 384360 X. This Second Edition updates developments in all areas of this field. The sections on inlets, sampling procedures, column selection, column maintenance and applications have been ex- panded. A number of phenomena, such as the complex inter-relationships of carrier gas choice and velocity, column characteristics, tempera- ture programme parameters on analysis times and separation efficiencies have been discussed in plain terms.New sections concerned with instrument conversion and special analytical methods have been added. The manufacture, care and use of conventional glass capillary columns and of new fused silica capillaries are covered. The Development and Use of Fume Cup- boards, Fume Hoods and Ventilated Safety Enclosures for Laboratories. Proceedings of a Symposium organised by the Labora- tory of the Government Chemist, 22 March, 1979, at Church House, Westminster, London, SW1. Pp. vi + 157. Laboratory of the Government Chemist. 1979. Gratis. The facet emphasised in this Symposium was safety, the conduct of potentially dangerous operations with confidence and with safety. Invitations to participate in the Symposium were limited to government laboratories and allied organisations (including University Chem- istry Departments) and those directly interested in the construction and maintenance and opera- tion of these.The papers presented were: Work in the UK on the Development of Fume Cupboards and Ventilated Enclosures ; Health and Safety Legislation and its Implications; Fume Cup- board Development for the Government Service ; The Use of Fume Cupboards and Related Items in the Chemistry Laboratory ; The Containment Performance of Fume Cupboards; The Aero- dynamics of Fume Cupboards ; Laminar Down- flow Cabinets-Some TJnseen Hazards ; The Open Bench Environment ; An Enclosed Cabinet System for the Handling of Highly Infective Agents; Fume Cupboards and Their Extraction Systems in Analytical Chemistry ; and The Discharge of Fume Cupboard Effluent into the Atmosphere.Chemical Industry Directory 1980. Pp. vi + 418. Benn Publications Limited. 1980. Price A25. ISBN 0 510 48712 2. Aquametry. Part 111. A Treatise on Methods for the Determination of Water. Second Edition. John Mitchell, Jr. and Donald Milton Smith. Chemical Analysis, Volume 5. Pp. xviii + 851.. John Wiley. 1980. Price k41. ISBN 0 471 02266 7. This third volume of the newly updated treatise on water measurement, “Aquametry,” com- pletes a definitive examination of current methods for determining water in liquids, solids and gases. Part I11 focuses on the Karl Fischer reagent (KFR), a technique unique in its ability to determine water a t concentrations ranging from fractional parts per million to essentially 100%.The authors provide a new critical assessment of the many variations of KFR preparation, includ- ing stabilised compositions, macro- and micro- techniques for analysis, new developments in electrometric methods for determining the end- point, and precautions in sampling and titration to assure high precision and accuracy. The book then reviews applications and limitations of the reagent in analyses of a wide variety of organic and inorganic species ranging from high purity compounds to technical grade commercial materials. Novel non-acidimetric procedures for determ- ining several classes of organic and inorganic compounds are described. The Index includes the major categories in Parts I and I1 to aid readers in choosing the methods best suited to their needs.Trace Chemistry of Aqueous Solutions. General Chemistry and Radiochemistry. Petr BeneS and Vladimir Majer. Topics in Inorganic and General Chemistry, Monograplz 18. Pp. 252. Elsevier. 1980. Price $56; Dfl115. The two main sections are (a) Traces in Homo- geneous and Microheterogeneous Aqueous Systems and (b) Traces in Macroheterogeneous. Systems : Aqueous Solution-Solid Phase. The former section is divided into (i) The Ionic and Molecular State of Traces and (ii) The Colloidal State of Traces, and the latter section into (i) Nature, Classification and Importance of Distri- bution Phenomena, (ii) Distribution of Traces between a Solution and a Solid Phase in Statu Nascendi and (iii) Distribution of Traces Between a Solution and a Pre-formed Solid Phase. ISBN 0 444 99798 9.December, 1980 PUBLICATIONS RECEIVED 553 in this section were presented as Review Papers a t an IAEA Symposium on Nuclear Activation Techniques in the Life Sciences, held in Vienna in 1978) and Analytical Quality Control.Gas Chromatography with Glass Capillary Columns. Second Edition. Walter Jennings. Fp. xiv + 320. Academic Press. 1980. Price $25. ISBN 0 12 384360 X. This Second Edition updates developments in all areas of this field. The sections on inlets, sampling procedures, column selection, column maintenance and applications have been ex- panded. A number of phenomena, such as the complex inter-relationships of carrier gas choice and velocity, column characteristics, tempera- ture programme parameters on analysis times and separation efficiencies have been discussed in plain terms.New sections concerned with instrument conversion and special analytical methods have been added. The manufacture, care and use of conventional glass capillary columns and of new fused silica capillaries are covered. The Development and Use of Fume Cup- boards, Fume Hoods and Ventilated Safety Enclosures for Laboratories. Proceedings of a Symposium organised by the Labora- tory of the Government Chemist, 22 March, 1979, at Church House, Westminster, London, SW1. Pp. vi + 157. Laboratory of the Government Chemist. 1979. Gratis. The facet emphasised in this Symposium was safety, the conduct of potentially dangerous operations with confidence and with safety. Invitations to participate in the Symposium were limited to government laboratories and allied organisations (including University Chem- istry Departments) and those directly interested in the construction and maintenance and opera- tion of these.The papers presented were: Work in the UK on the Development of Fume Cupboards and Ventilated Enclosures ; Health and Safety Legislation and its Implications; Fume Cup- board Development for the Government Service ; The Use of Fume Cupboards and Related Items in the Chemistry Laboratory ; The Containment Performance of Fume Cupboards; The Aero- dynamics of Fume Cupboards ; Laminar Down- flow Cabinets-Some TJnseen Hazards ; The Open Bench Environment ; An Enclosed Cabinet System for the Handling of Highly Infective Agents; Fume Cupboards and Their Extraction Systems in Analytical Chemistry ; and The Discharge of Fume Cupboard Effluent into the Atmosphere.Chemical Industry Directory 1980. Pp. vi + 418. Benn Publications Limited. 1980. Price A25. ISBN 0 510 48712 2. Aquametry. Part 111. A Treatise on Methods for the Determination of Water. Second Edition. John Mitchell, Jr. and Donald Milton Smith. Chemical Analysis, Volume 5. Pp. xviii + 851.. John Wiley. 1980. Price k41. ISBN 0 471 02266 7. This third volume of the newly updated treatise on water measurement, “Aquametry,” com- pletes a definitive examination of current methods for determining water in liquids, solids and gases. Part I11 focuses on the Karl Fischer reagent (KFR), a technique unique in its ability to determine water a t concentrations ranging from fractional parts per million to essentially 100%.The authors provide a new critical assessment of the many variations of KFR preparation, includ- ing stabilised compositions, macro- and micro- techniques for analysis, new developments in electrometric methods for determining the end- point, and precautions in sampling and titration to assure high precision and accuracy. The book then reviews applications and limitations of the reagent in analyses of a wide variety of organic and inorganic species ranging from high purity compounds to technical grade commercial materials. Novel non-acidimetric procedures for determ- ining several classes of organic and inorganic compounds are described. The Index includes the major categories in Parts I and I1 to aid readers in choosing the methods best suited to their needs. Trace Chemistry of Aqueous Solutions. General Chemistry and Radiochemistry. Petr BeneS and Vladimir Majer. Topics in Inorganic and General Chemistry, Monograplz 18. Pp. 252. Elsevier. 1980. Price $56; Dfl115. The two main sections are (a) Traces in Homo- geneous and Microheterogeneous Aqueous Systems and (b) Traces in Macroheterogeneous. Systems : Aqueous Solution-Solid Phase. The former section is divided into (i) The Ionic and Molecular State of Traces and (ii) The Colloidal State of Traces, and the latter section into (i) Nature, Classification and Importance of Distri- bution Phenomena, (ii) Distribution of Traces between a Solution and a Solid Phase in Statu Nascendi and (iii) Distribution of Traces Between a Solution and a Pre-formed Solid Phase. ISBN 0 444 99798 9.December, 1980 CONFERENCES AND MEETINGS Progress in Drug Metabolism. Volume 4. Edited by J. W. Bridges and L. F. Chasseaud. Pp. xii + 335. John Wiley. 1980. Price i 2 5 . ISBN 0 471 27702 9. The five chapters in this book include “The Gas Chromatographic Analysis of Drugs in Biological Fluids,” by C. M. Kaye, and “The HPLC Measurement of Drugs in Biological Fluids,” by P. J . Meffin and J. 0. Miners. 555

ISSN:0144-557X    

DOI:10.1039/AP9801700551

出版商:RSC    

年代:1980

数据来源: RSC

摘要:

December, 1980 CONFERENCES AND MEETINGS 555 information can be obtained from the Conference and Courses Unit at Sira Institute Ltd., South Hill, Chislehurst, Kent, BR7 5EH. Conferences and Meetings Sampling Systems for On-line Process Measurement February 11 and 12, 1981, London This title covers two separate seminars that will be jointly sponsored by Sira Institute Ltd. and the Warren Spring Laboratory. They will be held at the Sudbury Conference Hall, London, E.C.l. The first seminar, on Liquids and Gases, is expected to include the following papers. Scene-setting paper, A. Verdin ; “Off-line Sampling of Two-phase Mixtures,” P. F. Drake; “Reliability and Maintenance,” C. Everaerts ; “Sampling of Particulate Matter in the Steam/ Water Circuit of Turbine Generators,” G. A.Fitchett ; “High-pressure, High-temperature Systems,” T. Hibdige; “Sampling for Flue Gas Analysis and Boiler Control,” J. Laxton ; “Basic Principles of Sampling System Design, and Some Cautionary Case Histories,” C. W. Munday; and “Use of Inert Gases for Purging in Tankers and Bulk Carriers,” P. Parnell. The second seminar, on Slurries, Pastes and Solids, is expected to include the following papers. Scene-setting paper, speaker to be announced ; “In-stream zlersus On-line Samp- ling,” P. Ball; “The Automatic On-line Samp- ling of Solids for Analysis,” B. J. Hulley; “Sample Handling for On-stream Determination of Carbon in Pulverised Fuel Ash,” R. Kempster ; “Sampling Systems for Routine Testing of Plastics Powders and Granules,” R. C. Moore; ‘Samplex-sampling Systems for a Continuous Sample Flow for On-line and Laboratory Measurements in the Pulp and Paper Industry,” M.Myrttinen and T. Paakkonen; and “Hand- ling Non-Newtonian Pastes and Slurries,” I. Stewart. A literature display describing equipment commercially available for sampling systems will be associated with the seminars. Further Official Methods of Analysis and Stability Testing of New Drugs Maah 25 and 26, 1981, Cambridge The Chromatography and Electrophoresis and Joint Pharmaceutical Analysis Groups and East Anglia Region of the Analytical Division and the Eastern England Region of the Industrial Division of the RSC are to organise a Joint Symposium on the above topic at New Hall, Cambridge University. The speakers will be F. Bailey, J. Tillman, A. F. Fell, R.G. Hopkins, B. T. Croll, A. G. Croft, G. M. Telling and A. Hobson-Frohock. In addition, there will be a session on “The Work of the AMC” with three speakers. For further details contact Dr. D. Simpson, Analysis for Industry, Factories 2/3, Bosworth House, High Street, Thorpe-le-Soken, Essex, C016 OEA. Metallurgical Coatings April 6-10, 1981, S a n Francisco, USA This International Conference will be held at the Jack Tar Hotel. The sessions will be as follows : Metals and Alloy Coatings ; Solar Thermal Coatings ; Characterisation of Coatings and Defects; Metallurgical Aspects of Micro- electronics ; Metallurgical Aspects of Integrated Optics ; Refractory Compound Coatings; Tri- bological Coatings; Coatings for Use at High Temperature ; Coatings for Radiation Environ- ments ; Surface Modification by Directed Energy Sources ; Thermal Spraying Techniques and Applications ; Plasma Anodisation Techniques ; Plasma Polymerisation Coating Technology ; Coatings for Batteries and Fuel Cells; Amor- phous Films ; In Situ Coating Inspection and Repair of Large Engineering Components.The Conference Chairman is Professor R. F. Bunshah, University of California, 6532 Boelter Hall, Los Angeles, Ca. 90024, USA. RSC Annual Chemical Congress The Annual Congress of The Royal Society of Chemistry is to be held at the University of Surrey. The Analytical Division will be arranging a symposium entitled “Matrix and Sensitivity Problems in Analysis” in which the lectures will be given by W. L. Budde, N. T. Crosby, D. N. Hume, J. S. Leahy, E.McKerrell, J. N. Miller, A. C. Moffat, B. Scales, C. Scott, T. Stijve, G. M. Telling and A. P. Woodbridge. April 7-9, 1981, Guildford I556 CONFERENCES AND MEETINGS Anal. Proc The Theophilus Redwood Lecture will be delivered by Prof. J. D. Winefordner. Further information (the first circular) is available from Dr. J. F. Gibson, The Royal Society of Chemistry, Burlington House, Piccadilly, London, W 1V OBN. Anglo-Dutch Symposium on Quantitative Organic Analysis A p r i l 22-24, 198 1, Noordwijkevhout, T h e Nethev- lawds The above Symposium will be organised by the ,Analytical Chemistry Section of the Royal Netherlands Chemical Society and the Analytical Division of the Royal Society of Chemistry. The plenary lectures will be as follows : “Develop- ment and Evaluation of Selected Assays for Drugs and Drug Metabolites in Biological Materials,” by Professor A.de Leenheer ; “The Analysis of Additives and Residues in Plastics Materials,” by Mr. D. C. M. Squirrell; “Monitor- ing Exposure to Toxic Gases in Work Place Atmospheres,” by Mr. D. T. Coker; “The Com- puter Revolution and its Impact on Quantitative Organic Analysis,” by Professor D. Betteridge ; and “Various Applications of Functional Group Analysis,” by Mr. D. van Houwelingen. There will also be three keynote lectures: “Aspects of the Analysis of Drugs and Drug Metabolites,” by Dr. G. G. Skellern; “The Analysis of Polymers and Resins and Related Additives,” by Dr. J. Bax; and “Aspects of Quantitative Analysis of Organic Vapours in Work Place Atmospheres,” by Dr.J. P. C. M. van Dongen. For further details contact Dr. B. Griepink, Secretary of the Analytical Chemistry Section, Royal Netherlands Chemical Society, c/o Analytical Chemistry Laboratory, Croesestraat 77A, 3522 AD Utrecht, The Xetherlands. Detectors in Chromatography April 28, 1981, Southend The Chromatography and Electrophoresis Group of the RSC will organise a symposium with this title a t the College of Technology. The speakers will be R. M. Smith, K. S. Webb, A. D. Jones, D. J . Malcolme-Lawes and A. Townshend. For further details contact Dr. D. Simpson, Analysis for Industry, Factories 2/3, Bosworth House, High Street, Thorpe-le-Soken, Essex, C016 OEA. 18th Annual Short Summer Course in X-ray Spectrometry June 1-12, 1981, Albany, N.Y., USA This course will be offered a t the State Univers- ity of New York a t Albany.The course will be instructional and will develop the basic theory and techniques, starting from elementary principles. No previous knowledge or experience is required. The first week will cover basic principles, techniques and practical applications, and the second week will continue with further fundamentals and practical applications. Both weeks will illustrate and employ equally the wavelength-dispersive and energy-dispersive methods. Emphasis in the second week will be placed on advanced principles and techniques and absorption-enhancement corrections by several procedures including mathematical methods, computer calculations and computer automation of modern X-ray spectrometers. Equal time will be devoted to lectures and laboratory-problem solving sessions.Registra- tion may be made for one week either a t a fee of $650.00, or for the entire two-week session a t a registration fee of $1 200.00, payable in US dollars drawn on a US bank. For further information please communicate with : Professor Henry Chessin, State University of New York at Albany, Department of Physics, 1400 IYashing- ton Avenue, Albany, New York 12222, USA. 4th International Symposium on Affinity Chromatography and Related Techniques June 22-26, 1981, Eindhoven, The Nethevlands The meeting will be held a t the De Koningshof Conference Centre, Veldhoven, and will be divided into three parts : Theoretical Aspects; Polymeric Matrices and Ligand Immobilisation ; and Applications. An exhibition of scientific equipment, laboratory reagents and scientific books will be held in conjunction with the symposium.Proceedings will be published. For information write to Mr. P. J. M. Toll, Faculty of Science, Katholieke Universiteit, Toernooiveld, 6525 Ed Nijmegen, The Netherlands. Nuclear Magnetic Resonance July 12-17, 1981, Exeter This meeting will be organised by The Royal Society of Chemistry and held at the University. It will be composed of eight consecutive sym- posia : 1, Multinuclear Magnetic Resonance : Spin-$ Nuclei; 2, Intact Biological Systems; 3, Heterogeneous Systems ; 4, Multinuclear Mag- netic Resonance : Quadrupolar Nuclei ; 5, Applicable Theory ; 6, Dynamic Processes ; 7 and 8, Techniques. There will also be two poster sessions. A full programme of evening socialDecember, 1980 CONFERENCES events is being planned, together with a Wednes- day afternoon event and a ladies’ programme. For further information contact Dr.J. F. Gibson, Secretary (Scientific), The Royal Society of Chemistry, Burlington House, London, WlV OBN. IUPAC International Symposium : The Harmonisation of Collaborative Analytical Studies August 20-21, 1981, Helsinki, Finland This international symposium, to be held a t the Hotel Kalastaj atorppa, has been organised jointly by the Analytical Chemistry, Applied Chemistry and Clinical Chemistry Divisions of IUPAC in recognition of the importance of such studies in many areas of everyday life, ranging from health and environmental standards and control to all aspects of industry and commerce.It will be of direct interest to organisations that sponsor collaborative analytical studies and to those who participate in them. The interest is both national and international. Agreed analytical methods, standardised in this manner, are among the most important end-products of chemical investigation, teaching and research. There is now considerable interest in avoiding duplication and recognising a sound basis for the comparison of two or more methods, each of which has been validated by collaborative study. The symposium will consider the role and design of collaborative studies, with special reference to analytical parameters which are important in the validation and intercomparison of methods (e. g., accuracy, precision, reproduci- bility), and the definition and practical inter- pretation of such parameters.The discussion will relate both to national and international aspects of validation, the avoidance (or minimisation) of duplication of effort and the basic objectives in the choice of analytical criteria. National and international organisations interested in the development and use of stand- ard methods are invited to send representatives to the symposium, a selection of the papers presented to which will be published subse- quently in the IUPAC journal Pure and Applied Chemistry. Individuals with an interest in the field are, equally, invited to attend. Contribu- tions are invited from participating organisa- tions or their representatives. A preliminary registration form is available from Dr.G. Svehla, Department of Chemistry, The Queen’s University of Belfast, Belfast, BT5 9AT, Northern Ireland, from whom further information can be obtained. AND MEETINGS 557 Sixth Australian Symposium on Analytical Chemistry August 23-28, 1981, Canberra, Australia The symposium will be organised by the Analyti- cal Chemistry Division of the Royal Australian Chemical Institute and held a t the Australian National University. Papers and posters are invited in any branch of analytical chemistry and sessions are planned to cover chromato- graphy, education, electrochemical analysis, food analysis, instrumentation, mineral analysis, pharmaceutical analysis, plasma spectroscopy and trace analysis. A social programme will be arranged. Correspondence concerning the symposium should be addressed to the Symposium Secretary, 6th Analytical Chemistry Symposium, P.O.Box 1397, Canberra City, ACT 2601, Australia. International Conference on Mercury Hazards in Dental Practice September 2-4, 1981, Glasgow The topics to be discussed will include epi- demiology of mercury poisoning, methylation of mercury, toxicology, measurement and monitoring and control and prevention. Further details are available from Professor J. M. A. Lenihan, West of Scotland Health Boards, Department of Clinical Physics and Bio- Engineering, 11 West Graham Street, Glasgow, G4 9LF. 9th International Conference on Atomic Spectroscopy and XXII Colloquium Spectroscopicurn Internationale September 4-8, 1981, Tokyo, Japan The 1981 version of this conference will be held a t Sophia University, with accommodation a t The New Otani Hotel.It is to be organised by the Japan Society for Analytical Chemistry in co-operation with the Spectroscopical Society of Japan. Sessions will be held on plasma emission spectroscopy, atomic absorption spectroscopy, atomic fluorescence and scattering spectroscopy, Fourier transform spectroscopy, laser spectros- copy, computers in spectroscopy, microbeam and surface analyses, spectroscopy for chemical state analyses and applications to life science. Invited speakers who have agreed to attend include L. S. Birks, P. W. J. M. Boumans, C. L. Chakrabarti, K. Dittrich, L. de Galan, V. A. Fassel, K. Fuwa, K. F. J. Heinrich, G. M. Hieftje, G. Horlick, P. N. Keliher, G. F. Kirk- bright, K. Kohra, s. R. Koirtyohann, B.V. L’vov, R. J. MacDonald, A. Meisel, S. Minami, S. Nagakura, J. M. Ottaway, T. C. Rains, J.558 COURSE Anal. PYOC. Robin, A. Rosencwaig, I. RubeSka, G. A. Somorjai, J. C. Van Loon, A. Walsh, J. P. Walters, T. S. West, J. D. Winefordner and Zeng Yun-E. Further details on all aspects of the conference can be obtained from the Conference Secretariat, The Organizing Committee 9th ICAS and XXII CSI, c/o The Japan Society for Analytical Chemistry, Gotanda Sanhaitsu, 26-2 Nishigot- anda 1-chome, Shinagawa-ku, Tokyo 141, Japan. 12th Annual Symposium on the Analytical Chemistry of Pollutants April 14-16, 1982, Amsterdam, The Netherlands This symposium is to be held at The Free University, de Boelelaan 1105, 1081 HV Amsterdam, The Netherlands. The scientific programme consists of invited plenary lectures and invited and submitted research lectures and poster presentations covering the whole field of environmental analytical chemistry.As has been customary in previous symposia, one day will be devoted to the following interdisciplinary topics : Analytical Chemistry and Water Chem- istry ; and Analytical Chemistry and Mutageni- city (with special emphasis on water pollutants). Those interested in giving a research lecture (20 min including discussion) or a poster presenta- tion should send an abstract of no more than 200 words to the Congress Office by March 15, 1981. Further information can be obtained from Professor Dr. R. W. Frei, Congress Office, 12th Annual Symposium on the Analytical Chemistry of Pollutants, Congress Bureau, Vrij e Universi- teit, P.O. Box 7161, 1007 MC Amsterdam, The Netherlands. 9th International Mass Spectrometry Conference August 30-Se+teunber 3, 1982, Vienna, Austria The conference will take place in the castle “Hofburg” in Vienna. It will be organised by the Austrian Mass Spectrometry Group, the Austrian Society for Microchemistry and Analytical Chemistry, the Austrian Chemical Society and the Institute of Analytical Chem- istry of the University of Vienna in co-operation with an International Scientific Committee. The scientific programme will cover all aspects of mass spectrometry and will consist of invited lectures, submitted papers and posters. Also, an extensive commercial exhibition will be arranged. Further information is available from the secretariat of the conference : Interconvention, P.O. Box 105, A-1014 Vienna, Austria.

ISSN:0144-557X    

DOI:10.1039/AP9801700555

出版商:RSC    

年代:1980

数据来源: RSC