Abstract A radially compressed column, packed with micro particle, reversed phase (C 18) material, was used to study HPLC with cetrimide containing eluents. The amount of cetrimide adsorbed onto the stationary phase was measured; not the number of available adsorption sites, but rather the presence of micelles in the eluent appears to be the limiting factor for the uptake of cetrimide from the eluent. The capacity factors, κ′, of several - mainly acidic - compounds were determined in this system, with varying pH and cetrimide concentration of the eluent (methanol-water, 50% w/w). The results obtained upon changing the pH of the eluent can not all be explained with an ion-exchange (or ion-pair) model. Upon increasing the cetrimide concentration, maximum values in κ′ are reached at about the critical micelle concentration (cmc) of cetrimide in the eluent. The results of conductimetric experiments suggested, that the decrease of κ′ at cetrimide concentrations above this CMC, observed for most of the compound...

s of papers DETERMINATION OF THE ANTITUMOR AGENT ADRIAMYCIN IN RAT TISSUE W.J. van Oort, B. Kerkhofs and A. van Dijk Adriamycin is an an thracycl ine der iva t ive c u r r e n t l y in widespread clinical use . It is a ve ry act ive d r u g in sev eral types of tumors as small cell lung cancer and breas t cancer . Cardiac toxic i ty is one of i ts most severe toxic side effects . T a r g e t t i n g adriamycin by encapsulat ion in liposomes, it is necessa ry to inves t iga t e in pharmacological exper i ments the d i s t r ibu t ion of adriamycin in plasma and s eve r al t ypes of t i s sue of r a t s . In addit ion to the determination of adriamycin in plasma (1), a t i s sue sample p re t reatment is p re sen ted , s t a r t i n g by f r eeze -d ry ing the t i s sues , degenera t ion of the cells by addit ion of AgNO.~, shak ing for 10 minutes , ad jus t ing the pH at 8.9 and e~t r a c t i n g by ch loroforml -heptanol (1+1, v / v ) . The foliowing s t eps are ident ical to the descr ibed HPLC method (1). Prel iminary d i s t r ibu t ion p a t t e r n s obtained by adminis t ration of free adriamycin and in two types of liposomes are p re sen ted . (1) A.M.B. Bots, W.J. van Oort , J. Noordhoek, A. van Dijk, S.W. Klein, Q.G.C.M. van Hoesel, J . ChromatoRT., accepted Pharmaceutical Labora tory , Cather i jnes ingel 60, Utrecht Academic Hospital, Univers i ty of Utrecht , Cathar i jnes ingel 101, Utrecht , The Nether lands THE BEHAVIOUR OF INJECTED SALTS IN REVERSED PHASE SYSTEMS -IMPLICATIONS FOR THE DETERMINATION OF THE HOLD-UP TIME O.A.G.J. van derHouwen, A. Hulsh~ff and A.W.M. Indemans The interaction of salts of alkalimetals with reversed phase columns was studied by chromatographic means. For the interpretation the chromatograms have been compared with those obtained on non-modified silicagel under the same conditions. The results indicate that on injection of these salts three peaks are obtained. These peaks are proven to be caused by the injected anions, the injected cations and buffer concentration pulse respectively. The retention of the anion peak and the buffer concentration pulse peak are explained by charge exclusion and well known concentration pulse propagation concepts. The implication of these phenomena for the use of salts for the ~etermination of the column void volume are discussed. Pharmaceutical Laboratory, Department of Analytical Pharmacy, State University of Utrecht, Catharijnesingel 60, 3511GH Utrecht, The Netherlands r98 Vol. 4 I982 Pharmaceutisch Weekblad Scientific Edition DETERMINATION OF THEOPHYLLINE IN SERUM AND ITS BINDING TO SERUM PROTEINS WITH ISOTACHOPHORESIS J.C.Reijenga, A.Gaykema, F.E.P.Mikkers ............................................... The concentration of free theophylline in serum was determined with isotachophoresis in the therapeutical range 0-20 mg/l. Serum was ultrafiltrated , the filtrate diluted and directly injected. The bidding of the drug to total pool serum proteins, albumin and orosomucoid was determined as 55, 44 and 12% respectivily. The effect of different parameters during filtration was investigated. Laboratory for Instrumental Analysis, Eindhoven University of Technology, P.O.Box 513 Eindhoven (The Netherlands) Differential Pulse I~larographlc 0uantitation of the Vasodilator Nitroprusside in Human Serum. O.R.Leeuwenkampf H.Jousma, E.J. van der Mark, A.Bult Initially the following procedure for serum analysis was employed: protein elimination with i.~4 perchloric acid (i : 4), treatment with active charcoal, centrifugation, membranefiltration and analysis of the resulting serum solution with high performance differential pulse pularography. However, the peaks were considerably lower and distorted in comparison with IM perchloric acid solutions of nitroprusside due to the formation of nitroprusside complexes with copper and other metals present in serum. After elimination of the proteins and the metals with 1.2M perchloric acid containing 1 mg/ml ferrocyanide, the peaks have the height expected in case of 100% recovery. A linear calibration curve was obtained for spiked serum. Detection limit: 15 ng/ml. Gorlaeus Laboratories, dept. of Pharmaceutical Analysis and Analytical Chemistry, subfaculty of Pharmacy, Leiden, The Netherlands. DATA HANDLING IN ISOTACHOPHORESIS J.C.Reijenga, W.van Iersel ............................................... The properties of the universal detector signal in isotachophoresis prohibit the use of standard chromatographic integration procedures. Therefor a computerized conversion is applied to auto~ati cally monitor the signal and give both qualitative and quantitative inforn~tion, so that full use is made of the linear dynamic range (I0000) of the separation method. The method is illustrated with the analysis of oxalate, glycolate, citrate and phosphate in urine. Several data reduction procedures can be used in this respect. Laboratory for Instrumental Analysis, Eindhoven Universf~:y of Technology, P.O.Box 513,Eindhoven (The Netherlands) Title: MULTI-CHANNEL U.V.DETECTION AS AN AID FOR PURITY CONTROL USING HPLC.

S OF PAPERS SUBMITTED FOR PRESENTATION TO THIS SOCIETY The following papers have been submitted to the Secretary and the Associate Secretaries of the Society for presentation at meetings of the Society. They are numbered serially throughout this volume. Cross-references to them in the reports of the meetings will give the number of this volume, the number of this issue, and the serial number of the abstract. 192. Dr. A. E. Ross: On criteria for universality of ternary quadratic forms. Necessary and sufficient conditions are derived in terms of generic characters that an indefinite ternary quadratic form, classic or non-classic, represent all positive and all negative integers. Such a form must represent zero properly. The theorem for classic ternary quadratic forms was proved by the author (Abstract, this Bulletin, vol. 36 (1930), p. 364) by use of a result of A. Meyer. This paper proves the theorem without use of the equivalence criterion of Meyer and similarly derives the theorem for non-classic forms. Theorem 1. Let ƒ be a properly primitive indefinite classic ternary quadratic form with reciprocal F and determinant D. The necessary and sufficient conditions tha t ƒ be universal are: D is odd or double an odd integer, 0 = ± 1, and (F/p) = ( — ü/p) for every odd prime p dividing A — D. Theorem 2. Let / i be a primitive indefinite non-classic ternary quadratic form. Consider an improperly primitive form / = 2 / i with reciprocal F. Then / i is universal if and only if f—2fi satisfies the following conditions: Ü—±\, the characters of ƒ are (_l)(F-D/2 = (_i)(-r-i)/2> (F/p)^(-ü/p) for every odd prime p dividing A — D, and, in case 4 divides A, also ( — l ) ^ _ ) / = 1. These theorems serve as a practical test for determining whether a given ternary quadratic form is or is not universal. (Received July 19, 1932.) 193. Dr. C. F. Luther: Concerning primitive groups of class u. In this paper are proved three general theorems concerning the degree and class of multiply transitive groups. The first gives an upper limit to the degree of a substitution group of class u that contains a substitution of order two and degree w+e (e a positive integer), and is more than £ 1 + ^ 2 + ^ 3 + • • • Pr times transitive, where pi, pi, p%, • • • , pr are distinct odd prime numbers and r>l. I t is u>n-€-(n+2ep1p2 • • • pr)/'((pi-1)(pi-1) • • • (pr~l)) if e/u is small. Limits are also given for 2, 3, 5, 6, 7, and more than p (a prime)-ply transitive groups. The second theorem gives an upper limit to the degree of a triply transitive group of class u(>3) tha t contains a substitution of degree u-j-e (e a positive integer) and of order p (p an odd prime). The third theorem gives an upper limit to the degree of a doubly transitive group of class u that contains a substitution of degree u-\-e (e a positive integer) and of prime order p (p an odd prime). (Received July 20, 1932.) License or copyright restrictions may apply to redistribution; see https://www.ams.org/journal-terms-of-use 1932.] ABSTRACTS OF PAPERS 633 194. Dr. Rothwell Stephens: Continuous transformations of finite spaces. The following problem is considered: Given a finite space P and a knowledge of the existence or denial of eight fundamental properties in Pt what possible combinations of these properties can occur in the biunivocal continuous transforms of P. The eight properties considered are those studied by D. McCoy (Tôhoku Mathematical Journal, vol. 33 (1920), pp. 89-116). Solutions are given for several particular types of spaces. In addition several extraneous theorems are proved. For every finite space of type P4 of more than one element, the derived set of any set E is given by K(E) —E-\-p. Every finite space of type P48 is homogeneous, and for every set E, K(E) —E. (Received July 21, 1932.) 195. Professor H. L. Miller: On the summability of double Fourier series. Those writers who have applied Cesàro's method of summing series to the study of the Fourier development of a function of two variables have considered only integral orders of summability. This investigation considers nonintegral orders in Cesàro's method of summing the double Fourier series. The Fourier development of a function of two variables, f(x, y), integrable (L) in the region ( — i r ^ x ^ w , —ir^y^Tr), will be summable (Ck), k>0, to the value of the function at any point within the region at which the function is continuous, provided the function remains finite in some cross-neighborhood associated with the point. At a point of discontinuity (xi, y{) which lies on a straight line or curve containing all other points of discontinuity in the neighborhood of (xh yi)y the development of a function satisfying the same restrictions will be summable (Ck), k>0, to a value half way between the limiting values of the function as the point (xi, y{) is approached from either side of the line of discontinuity, provided these limiting values exist, except when the tangent to the curve of discontinuity at (xh y{) is parallel to an axis. In this case the series will be restrictedly summable. (Received July 22, 1932.) 196. Professor G. S. Bruton: Certain aspects of the theory of equations f or a pair of matrices. Two square matrices A and B with characteristic values ai and pi are said to have property P 3 if any polynomial, f (A, B), in A and B has its characteristic values among the numbers f (at, pi). Sufficient conditions that A and B have property Pz are found in terms of common invariant direction. Necessary and sufficient conditions are found for the 1st, 2d, and 3d order cases. Special studies are made where ƒ (A, B) is limited to being A-\-B or AB. (Received July 22, 1932.) 197. Professor M. H. Ingraham: A study of certain related pairs of square matrices. Dr. G. S. Bruton studied matrices A and B with characteristic values ai and pi and such that f (A, B) had characteristics ƒ (a t-, pi). Further necessary condiLicense or copyright restrictions may apply to redistribution; see https://www.ams.org/journal-terms-of-use 634 ABSTRACTS OF PAPERS [September, tions that two matrices A and B have this property are found and properties of such pairs are discussed. (Received July 22, 1932.) 198. Professor T. H. Hildebrandt and Dr. I. J. Schoenberg: On linear functional operations and the moment problem for a finite interval in one or several dimensions. In the first part of this paper it is shown that the Riesz theorem that a linear continuous functional operation on continuous functions is expressible in the form fofda, is deducible from and consequently equivalent to the Hausdorff theorem giving necessary and sufficient conditions that a function of bounded variation on a finite interval be determined by its moments fê-xda. This suggests in turn a simple direct proof of the Riesz theorem. The second part of the paper derives results related to Stieltjes integrals in two variables and applies them to the extension of the Hausdorff-Schoenberg results on the moment problem as well as the Riesz theorem in more than one variable. (Received July 22, 1932.) 199. Professor Glenn James: On Fermât s last theorem. This paper considers the Fermât equation x+y=zt z>y>x>0, for the so called first case and proves that z~y>cn where c is a certain function of x, y and z whose lower limit is 2. This work provides a simple, and what seems to be a new proof for the case n = 3, and suggests a point of at tack on the general problem. (Received July 23, 1932.) 200. Professor W. A. Manning: The degree and class of multiply transitive groups, I I I . Let n be the degree and u the class of a group of substitutions G that is neither alternating nor symmetric. The author has stated that if G is 5-ply transitive, n S 2u; if 6-ply transitive, n <5u/3 ; if 8-ply transitive, n<8u/5 ; and finally, if G is as much as 12 times transitive, n<3u/2. The proofs of the last two limits depend on the proposition that if in a / ( > 6 ) times transitive group, all the substitutions of degree u+eor less are of odd order, then n< 2u—4e — It + 13. This is used in combination with the limits found by Dr. C. F . Luther for the degree of /-ply transitive groups of class u in which there are substitutions of degree w+e of even order. (Received July 23, 1932.) 201. Mr. H. M. Bacon: An extension of a certain theorem of Kronecker, A certain theorem of Kronecker may be stated as follows: Constants ai, «2, as, • • • , otn being linearly independent while jui» M2, M3, • • • , Mn are arbitrarily given real numbers, there exists a real number t such that differences ajt—fjLj, ( j = l , 2, 3, • • • , w), are in absolute value and modulo 1 less than any arbitrarily preassigned number e. This paper shows that if € = 1/N is given, and the constants «i, «2, «3, • • • , an are not linearly independent, then the inequalities \otjt—VJ \

The influence of methyl-, hydroxy and amino substituents on the electrochemical behaviour of simple 1,4-naphtho-and 1,4-benzoquinones, model compounds of many quinoid antitumour agents, in aqueous media was studied. Significant changes in electrochemical behaviour were observed, potentially the result of a change in the electron density of the quinone moiety, pre- or post-protonation of substituents, hydrogen bond formation, tautomerization reactions and steric interactions between the quinone moiety and substituents. The information obtained was of benefit in the elucidation of the reduction mechanisms of quinoid antitumour agents such as aziridnylquinones and mitomycins.

Tobramycin (TBM) is an aminoglycoside antibiotic mostly used for infections caused by gram-positive although it can be indicated for some gram-negative organisms. Due to the absence of chromophore groups in TBM and its basicity, several different analytical approaches have been described in the literature for its quantification using HPLC that address specific aspects of its physical–chemical properties. Some methods are based on derivatization with chromophores and fluorescence molecules. Others apply techniques for detection such as mass spectrometry, electrochemical pulse detector, amperometric pulse detection, refractive index and evaporative light scattering. There are also approaches to the separation that include capillary and micellar electrophoresis. The applicability of different types of detectors in the quantification of TBM, along with its degradation products, are discussed to present advantages and disadvantages as well as the use of ion pairs and different stationary phases.

A sensitive and selective high-performance liquid chromatographic (HPLC) method for the determination of vinblastine and vincristine in plasma and urine is described. The drugs are isolated from 1.0 ml of the biological fluid with a solid-phase extraction column (Bond-Elut Diol®). The HPLC method was combined with electrochemical detection at +850 mV versus an Ag/AgCl reference electrode. The detection limit is 100 pg for vinblastine and 250 pg for vincristine with a signal-to-noise ratio of 3, which permits the determination of these compounds in biological fluids at the nanogram level. Evaluation of the isolation method revealed that the drug recoveries and the reproducibility of the extraction procedure depend on the batch number of the solid-phase extraction column used.

A comparison is made between the use of aluminium oxide and non-modified silica gel as cation-exchange materials for the separation of basic drugs (amines) with aqueous solvent mixtures. The retention behaviour of the amines is studied and appears to be controlled predominantly by the pH and the concentration and nature of the modifier; the nature and concentration of the competing ions and the buffer components of the mobile phase also exert some influence on the retention. Preparations with imidazoline and tetracycline derivatives have been analysed as examples of the application of these ion-exchange systems on non-modified silica gel and aluminium oxide in the analysis of pharmaceutical formulations.

This paper describes the pharmacokinetics of teniposide (VM-26) after being administered iv in high doses to eight cancer patients (maximum dose, 1.0 g/m2). VM-26 levels in plasma, urine, saliva, duodenal fluid, and cerebrospinal fluid were determined using high-performance liquid chromatography in combination with electrochemical detection. The plasma concentration-time curve of VM-26 showed a triphasic decay with a slow third phase in five patients, whereas in two patients the plasma concentration decay was biphasic. The plasma pharmacokinetics of VM-26 proved to be linear and could be fitted to a three-compartment model (five patients) and to a two-compartment model (two). The steady-state volume of distribution varied from 13.2 to 24.7 L/m2. The total-body clearance ranged from 5.84 to 10.18 ml/minute/m2. Low concentrations of VM-26 were found in saliva, duodenal fluid, cerebrospinal fluid, and urine. Excretion of unchanged VM-26 into the urine varied from 8.8% to 13.9% of the administered dose. No glucuronide of VM-26 could be detected in plasma or other biological fluid.

The concept of bioreductive alkylation as a mechanism of action of quinone-containing anticancer agents was investigated, using electrochemical techniques. According to this concept, an electrochemical step (reduction of the quinone ring) is followed by one or more chemical steps, leading to formation of the actual alkylating species. The proper use of electrochemical analysis of potential bioreductive alkylating quinones in the design of new analogs is limited. Up to now, the only electrochemical parameter frequently used in structure-activity relationship studies, is the half-wave potential of the quinone reduction. However, reliable information can only be obtained from the found value of this parameter when the reduction mechanism has been elucidated. Furthermore, it only gives information about the first step of the model. More detailed electrochemical analysis of potential bioreductive alkylating quinones, in combination with a biological evaluation, is required to gain more insight in their mechanism of action and to yield quantitative information about substituent effects on both the electrochemical and the chemical step(s) of the model. Results of such studies of a series of aziridinylquinones indicate, that the biological activity in vitro is correlated with the ease of protonation of the aziridines after quinone reduction, which is in accordance with the concept of bioreductive activation. No correlation with the ease of protonation of the aziridines prior to quinone reduction or with the quinone reduction step itself can be found.

The disposition of pentamethylmelamine (PMM) was studied in the male Wistar rat. PMM (5 mg/kg) was administered intraarterially, i.v. (5 and 10 mg/kg), via the portal vein, and into the duodenum to cannulated and unanesthetized rats (n greater than or equal to 4) via infusion. Parent compound and metabolites were quantified by gas chromatography. The areas under the plasma concentration-time curves of PMM after intraarterial and i.v. administration were equal and twice as large as the areas after portal vein and intraduodenal administration. This indicated insignificant lung metabolism for PMM; the low bioavailability of PMM when given via the portal vein or intraduodenally (in both cases, some 50% of an i.v. dose) was the result of presystemic metabolism in the liver. PMM was completely absorbed after intraduodenal administration, and no intestinal metabolism was observed. Linear kinetic behavior of i.v. PMM was observed in the 5- to 10-mg/kg dose range. The area under the plasma concentration-time curve of the first metabolite N2,N2,N4,N6-tetramethylmelamine was significantly greater when PMM was given via the portal vein or intraduodenally than when given intraarterially or i.v. This indicated either extrahepatic elimination/renal excretion of PMM or the existence of an additional metabolic pathway. However, experiments with adrenalectomized rats and rats with ligated blood flow to the kidneys did not alter the area for the first metabolite. These findings may be explained by the formation of unknown metabolites and/or reactive intermediates of PMM.

The reaction between to bramycin and 1-fluoro-2,4-dinitrobenzene was studied. This reaction shows a sharp pH optimum because, even under moderate alkaline conditions, derivatization of the aliphatic hydroxyl groups of tobramycin becomes an important side reaction. Phosphate and phthalate buffers also react with the derivatization reagent. Buffers comprising tertiary amines and hydrochloric acid are preferred.

The principles and applications of fluorescence detection and fluorescence introducing reagents and methods in HPLC are reviewed. The design and requirements for fluorescence detectors, flow cells and excitation sources and the conversion of non-fluorescent compounds into fluorescent products by pre-column and post-column derivatization reactions are discussed. For the applications the emphasis is on drug analysis, where possible in biological fluids (serum, urine, etc.). The last paragraphs are divided in a number of sections in which newly developed and some scarcely used reagents are mentioned shortly; a more complete treatment is given of the reagents and labels most frequently used in the derivatization of certain functional groups. In this discussion the methods of derivatization as well as the selectivity, stability, fluorescence behaviour of the reagents/labels and derivatives and the reaction conditions are included. An up-to-date survey of the applications of fluorescence detection in liquid chromatography (TABLE III, TABLE IV and TABLE V), ends this review paper.