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The paper chromatography of phenols (expecially phenolic acids), aglycones, glycosides and certain sugars is carried out with two new solvents. The authors have also examined these substances with the aid of paper electrophoresis and a borate buffer. A detection reagent for o -diphenols on paper is described.

Minimization of samples for infrared spectral measurement was attempted by the use of a Japanese folkcraft, hand-made paper, impregnated with liquid paraffin. By this method, it is possible to measure 0.1 mg. of a sample. This hand-made paper was also used for paper chromatography and the developed paper was submitted directly to infrared spectral measurement. Component of Penicillium islandicun and inorganic sulfate ion were determined by spectral measurement of their chromatogram.

An aqueous solution of sodium molybdate has been used as a specific reagent for the detection of phenolic compounds with o-dihydroxy groupings. The paper electrophoretic behaviour of phenolic compounds has been studied using a series of different buffers with pH values ranging from 5.2 to 10.0. o-Dihydroxy compounds can readily be detected by the use of ammonium molybdate buffer, and other structural features can be revealed by the choice of suitable electrolyte solutions. Paper partition chromatography of phenolic compounds using molybdate-treated paper in described.

The paper chromatography of vat and sulphur dyes, using reducing solvents at 80°, has been carried out, the dyes being separated as their leuco compounds. In both cases ascending paper chromatography was applied. The relation between dye constitution and chromatographic behaviour has been examined.

A new solvent system was given for the chromatography of food preservatives; benzoic acid, salicylic acid, p-hydroxybenzoic acid and it's esters, p-chlorobenzoic acid, sorbic acid, resorcinol and β-naphtol with the ascending method on Toyo Roshi No. 50 paper at 20-25°. Cyclohexanol-ammonium carbonate buffer gave results in 30 hours. Spots were detected by absorption when the paper was examined in ultraviolet light and with color reagcnts of the methyl red or diazo spray reagent. Rf values for this solvent were listed for these substances. Each substance was separated except in the case of mixture of benzoic acid and sorbic acid. However in this case, these two substances were identified by measuring spectrophotometrically the alcohol solution extracted from the part containing the substances on paper chromatogram.Horizontal paper electrophoresis on Toyo Roshi No. 51 paper for 1.5-2 hours at 20V per cm with buffer consisted of dimethylformamide, pyridine, acetic acid and water (pH 5.6) or of butanol, ammonia, acetic acid and water (pH 5.0) was useful. MNC (the ratio of the migration of the substances of New Coccine) and colors given with the methyl red, FeCl3 and diazo spray reagents were tabulated adding dehydroacetic acid to above substances. Data was also given for citric acid and tartaric acid.Sake (Japanese wine), soy etc., was acidified with HCl, extracted with ether, purified as ammonium-salts and submitted to paper chromatography and paper electrophoresis with similar technique.

A number of simple chromatographic applications are described in which sheet paraffin or other plastic film is used as an adhesive vapor barrier: 1. A funnel attached to a paraffin-enclosed narrow paper wick for spotting large volumes of sample solution as small spots of controlled size. 2. A funnel for the elution of a spot and its reapplication to another paper in a single operation. 3. A method for elution or chromatography which requires no jars or other rigid saturation chambers.

Energy calculations for manufacture of paper from logs and waste papers are reported based on data on energy use from the Canadian Census of Manufactures and estimates for collection and de-inking of waste. The energy requirement of the recycling technology explored exceeds the energy requirement of manufacture from logs by 33%. It appears that forest conservation through recycling has a fossil fuel cost.

Modified paper chromatography procedures for the analysis of fatty acids produced by anaerobic bacteria are described. Both ethylamine and hydroxylamine derivatives of fatty acids were prepared from inoculated anaerobic culture broth. The derivatives were spotted on chromatography paper and developed with appropriate solvents. Paper chromatography is a valuable alternative to gas liquid chromatography as an ancillary procedure in the identification of anaerobic bacteria in the clinical bacteriology laboratory.

The automated dithionite test for the detection of hemoglobin S has been modified for use with AutoAnalyzers having a filter paper read-out. This method was compared with results of hemoglobin electrophoresis in 425 individuals, of whom 58 were positive for hemoglobin S. The two methods were in complete agreement. The automated filter paper dithionite test is a rapid, economical, accurate method for detection of hemoglobin S and could be easily incorporated into currently available mass screening equipment.

An ascending chromatographic method was used to identify acidic mucopolysaccharides. On silica-gel-treated glass papers, a displacement type of chromatography with ethanol-salt solutions separated seven mucopolysaccharides. The compounds were located by charring with sulfuric acid or different staining technics. The method is convenient, reproducible, and rapid and can be used to identify small quantities of mucopolysaccharides in mixtures. Certain difficulties as and identification procedure were observed, however, and are discussed. Observations on glass-fiber papers are useful in studying different reactions of mucopolysaccharides with various salts and stains.

The design and manipulation of an apparatus for centrifugal acceleration of paper chromatography has been described. The main novel features consists of using filter paper strips instead of circular sheets and a constant flow solvent reservoir enclosed in the chromatography tank. Our arrangement is particularly suited for the quantitative analysis of materials labelled with radioactive isotopes of short half-lives and examples are given of the separation of some 82 Br- and 131 I-labelled substances. Advantages of the above method over circular paper chromatography are discussed.

The separations of steroid oestrogens achieved by paper chromatography have been reviewed with special reference to studied involving epimeric oestrogens and to oestrogens which have been isolated recently. Methods of detection of these compounds on paper chromatograms have also been discussed. The mobilities of thirty-four different oestrogens and their derivatives in many solvent systems have been tabulated. It is hoped that this review will be found useful in providing a collection of data from which separations of mixtures of oestrogens can be planned.

1. (1) The binding of acid dyes by denatured protein on filter paper has been investigated with special reference to Light Green. 2. (2) The uptake of dye, measured by elution, has been shown to vary according to the method of denaturation, the nature of the dye solvent (particularly the salt and and ethanol concentration) and finally with the duration and temperature of the subsequent washing procedure. 3. (3) Albumin and globulin do not behave in the same manner with different denaturing agents so that different methods give different albumin/globulin dye uptake ratios. 4. (4) The stoichiometric relationship between dye and protein is consistently nonlinear even at lower concentrations of protein on the paper and the same pattern can be demonstrated with protein on cellulose acetate membrane. It is suggested that this non-linear relationship at low protein densities is due to interference by the cellulose supporting medium and other anions. 5. (5) Intrinsic constants and the number of binding sites at infinite dye concentration have been measured under standardized conditions for Light Green, Brom-Cresol Green and Lissamine Green. The results have been compared and differences discussed. 6. (6) In the quantitative measurement of protein based on dye uptake, the importance of restricting the protein density on the paper is stressed.

A technique is described by which samples of tissue weighing from 1 to 10 mg can be frozen and ground and then stuck to paper chromatograms and the soluble compounds separated by normal solvent development. The advantages and special applications of this technique are discussed.

1. 1. A method for the detection of Jaffe-positive substances in human blood serum using the technique of paper partition chromatography is described. 2. 2. A partial solution of the problem of apparent creatinine has been furnished by identification of pyruvate as a second Jaffe-positive compound beside creatinine in a water-soluble fraction of acetone-filtrate of sera; this finding was confirmed polarographically as well as spectrophotometrically . Up till now the quantitative significance of this substance has not been considered in connection with this problem. 3. 3. After a few preliminary experiments, paper chromatography of the insoluble residue of acetone-filtrate of serum has been left open for further investigation. It seems likely that the final solution of the problem of apparent creatinine will cover both possibilities which were discussed in the introduction, irrespectively of whether the chromatographic investigation of the insoluble fraction might show the presence of some new unknown Jaffe-positive component.

A method is described for rapid analysis of phenylalanine and tyrosine using eluates of whole blood that has been impregnated into filter paper. This method employs semi-automated ion-exchange column chromatography (amino acid analyzer) and will yield reasonably accurate and consistent quantitative values for phenylalanine and tyrosine concentrations within two hours. The technique has significant value in rapidly confirming the presence of increased phenylalanine with or without a concomitant increase in tyrosine concentration in autoclaved or unautoclaved blood filter paper specimens. Thus phenylketonuria can be differentiated from hyperphenylalaninemia due to tyrosinemia within hours after the initial identification of an elevated phenylalanine concentration by a screening method such as the Guthrie test. In addition, further evaluations of phenylalanine and tyrosine concentrations can be facilitated in infants on whom only blood filter paper specimens are obtainable.

The effect of hemicellulose contents in pulp on beating rate and strength of paper made from it was studied.In this study two methods were used, one the adding method (adding isolated hemicellulose to the standard pulp) and the other the extracting method (removing hemicellulose stepwisely).In the former when the hemicellulose is isolated with drastic agents, chemical nature of hemicellulose is liable to change, so that it is impossible to say that the isolated hemicellulose will act in the same manner as it does in the original state. Therefore extracting method was employed in the present research.For the preparation of chlorite holocellulose, thin pieces of beech and aspen wood were treated with hot acidified sodium chlorite. These chlorite holocelluloses were treated with KOH solution under nitrogen gas, in order to minimize the oxidation of cellulose. For hemicellulose extraction, KOH solutions of various concentration (from 0.3 to 20%) were used in order to get a series of pulp in which the hemicellulose content was stepwisely arranged.The beating rate and the sheet strength of this series of pulp were examined. From the result of this study, it was concluded that there appeared to be the optimum hemicellulose content in pulp as for as the paper strength was concerned. Other observations were also made with regard to the result observed.

Paper chromatography is very helpful in the identification and comparison of antibiotics, but very few results from the application of this technique have been published so far. The awakening of interest in the search for antibiotics which are active against fungi pathogenic to plants and animals makes the investigator aware of the scant information that is available to help characterize such agents. A knowledge of the Rf values of the known antifungal agents would help advance the search for such therapeutic materials. Often these values, in conjunction with the biological inhibitory activities, are the only criteria by which one can determine whether the unknown agent appears to be a new material. When the Rf data are obtained for shake cultures, one can eliminate many previously described antibiotic-producing organisms from further study, thus allowing more research to be applied to antibiotics which are new and different. The studies reported in this paper should serve as a guide to the identification of some of the known antifungal antibiotics.

Examination of urinary product of rabbits administered with MHB, by filter paper chromatography, indicated that ureide, 3-OH-MHB, 3-keto-MHB, 3-keto-nor-MHB, and unknown compounds (MHB-M (VI)) were excreted. An excellent method for the separation of these compounds is the use of a buffered filter paper and butanol saturated with borate-NaOH buffer (pH 11). For detection of the located metabolites, 1% solution of sodium periodate and 1% solution of potassium permanganate were used and ultraviolet spectrophotometry was adopted for the characterization of barbiturates.

A simplified method for the quantitative determination of keto-acids by means of paper chromatography of their 2,4-dinitrophenylhydrazones is reported. 1. 1. The identification of these compounds on the filter paper is discussed, especially of those compounds which give more than one spot. 2. 2. The method is described in detail, espicially with regard to the pyruvic and α-ketoglutaric acid content in biological material. The recovery of these two keto-acids, starting from known aqueous solutions, averages 94%; on the basis of these results a standard curve can be drawn. According to this curve, recovery from deproteinized extracts of liver and blood averages 91.6% for pyruvic acid and 96.7% for α-ketoglutaric acid. 3. 3. Using this method the pyruvic and ketoglutaric acid content of rat blood, muscle, liver and brain is estimated.

Food preservatives—sorbic, benzoic, p-hydroxybenzoic acid and p-hydroxybenzoates—have been extracted from foods directly on to chromatographic paper with ethyl ether. Paper chromatographic separation of the extracted compounds was effected on the same paper. Methyl, ethyl and propyl p-hydroxybenzoates have been separated from each other during ether extraction and from the critical pair, the butyl and benzyl p-hydroxybenzoates, on a paper impregnated with sodium hydroxide. An examination was made of the reactions which distinguish sorbic and benzoic acids when both are present.

A technique has been developed by which complex mixtures of amines can be sepated by column chromatography on the ion exchange resins Amberlite CG-50 and Amberlite CG-120. Identification of amines is made on the basis of the speed of their elution from the resin column, as well as by subsequent paper chromatography of effluent fractions. The method can be applied to amines derived from various physiological fluids, and offers the possibility of isolating unidentified compounds for chemical characterization. Using the combined technique of ion exchange column chromatography followed by paper chromatography, 26 amines were identified in human urine. these were: methylamine, dimethylamine, ethylamine, ethanolamine, β-hydroxypropylamine, pyrrolidine, piperidine, putrescine, cadaverine, 2,2'-dithiobis(ethylamine), histamine, N-acetylhistamine, i -methylhistamine, p -hydroxybenzylamine, 3-methoxy-4-hydroxybenzylamine, p -tyramine, m -tyramine, octopamine, synephrine, 3-methoxytyramine, normetanephrine, metanephrine, tryptamine, serotonin, bufotenin, and kynuramine. An additional 20 unidentified bases, presumably amines, were found to be regularly excreted in urine. Evidence is presented supporting the identification of kynuramine, β-hydroxy-propylamine, putrescine, cadaverine, and 2,2'-dithiobis-(ethylamine). These 5 amines have not previously been considered to be constituents of normal human urine. It is suggested that study of the urinary excretion of amines may provide useful clues as to the mechanisms involved in various metabolic disorders, including those characterized by mental dysfunction.

Three products formed during the paper chromatography of glycine with water-saturated phenol as a solvent have been investigaetd. These unknowns were stable condensation products whose formation was dependent upon the presence of glycine ad formaldehyde. They were ninhydrin sensitive. Alkaline hydrolysis yielded glycine. For chromatographic practices, the amount of glycine or formaldehyde which may be converted into the condensation products will depend upon the amount of the other component in the mixture. Addition of 1% formaldehyde normally complexes all of the glycine. In usual chromatographic experiments part of the glycine may be complexed by residual formaldehyde present in chromatographic phenol. Glycine condensation may be minimized by redistilling the phenol solvent with aluminum turnings and sodium bicarbonate and by washing the filter paper with 1% oxalic acid.

A paper chromatographic system was developed for the resolution of mixtures of sulfated mucopolysaccharides. The effect of varying the ammonium formate buffer/isopropanol ratio in the developing solution on the RF values of heparin and chondroitin sulfate is presented. A complete separation of heparin from chondroitin sulfate was obtained with three solvent mixtures containing different proportions of buffer and isopropanol. One of these solvent mixtures, ammonium formate buffer-isopropanol (65:35) v/v, resolved commercial beef heparin into two principal components, on “fresh” paper. This system was used to compare beef heparin with mouse mast cell tumor heparin, rat heparin, sheep lung heparin, chondroitin sulfate, and β-heparin. A common metachromatic component with an RF of approximately 0.5 was0found in all of the heparin preparations chromatographed with this solvent mixture. “Aging” of the filter paper resulted in the loss of the ability of the buffer-isopropanol mixture (65:35) v/v to resolve the beef heparin. Increase of the buffer-isopropanol ratio to (70:30) v/v restored the resolution of the beef heparin on “aged” paper. Heparitin sulfate and α-heparin monosulfate were compared to beef heparin using the latter system.

A wide range of Δ 4 - and Δ 1 , 4 -3-ketosteroids, from weakly polar to strongly polar, may be chromatographed and detected on cellulose triaacetate paper, with benzene—methanol—water mixtures as developing solvents. Three systems have been worked out. In the first a one-phase mixture of benzene, methanol and water is used as the mobile phase and the paper is not accommodated. A “reversed phase” type of chromatogram is obtained, i.e. the more polar compounds have higher R F values than the less polar ones, as is usual for chromatography on acetylated paper. If, howerver, two-phase mixtures of benzene, methanol and water are used, as was the case in the other two methods, “reversed phases”, as well as “convetional” type chomatograms may be obtained. This depeds upon whether the lower or the upper layer is used as the mobile phase, the other layer being used for accommodation of the paper. Perhaps the first-mentioned method should, in contrast to the other two, be regarded as adsorption chromatography, rather than as partition chromatography. Well-reproducible R F values are obtained and, especially with the two-phase mixtures, compact spots are found. Detection methods and R F dta are given for various Δ 4 - and Δ 1 , 4 -3- ketosteroids, ranging from C 18 to C 27 compounds and including sex hormones and adrenoccortical hormones.

Publisher Summary This chapter discusses the principles of paper chromatography of antibiotics. It includes all steps of analysis beginning with the preparation of samples and papers, development techniques, detection methods, and means of documentation. Paper chromatography of the best-known groups of antibiotics is also presented in the chapter. General theoretical aspects of paper chromatography are not dealt with except for those that directly emerged from studies of antibiotics. The chapter concludes with a discussion of paper chromatographic classification and systematic analysis of antibiotics.

1. Low voltage paper electrophoresis in 0.05 M citrate buffer of pH 4.8, at a voltage of 200 V, for 312 hours, is a suitable method for determining adenine nucleotides in erythrocytes. 2. Before applying the sample to the paper it is necessary to remove the excess of deproteinisation agent. 3. By means of the method described 42.6 ± 14.5 mg% ATP and 10.7 ± 9.5 mg % ADP were determined in the erythrocytes.

Paper chromatography of pectin-methylesterase and polygalacturonase produced by Clostridium felsineum was achieved with buffers, buffered salt solutions and acetone-water mixture. No true separation of both enzymes was attained and they became inactivated when chromatographed on paper under appropriate conditions. The chromatographed enzyme preparation showed only pectin-methylesterase activity when the acetone-water solvent was used as the developing agent. The aforementioned enzyme occupied the lower part of the chromatograms on using the ascending technique. The inactivation of pectolytic enzymes was attributed to their denaturation by spreading in the form of a film on the paper chromatograms. The extent of solubility of these enzymes in the developing agent was found to be a limiting factor for their spreading on the chromatograms and, hence, their inactivation.

A critical review of the paper chromatography of Strychnos alkaloids has led to the following conclusions: 1. Paper chromatography is one of the most important methods of approach in elucidating the chemistry of Strychnos alkaloids. 2. Paper chromatography and electrophoresis, in the case of Strychnos tertiary alkaloids, are of great value for the rapid analytical identification of the various compounds. In the case of quaternary alkaloids these methods can give general information, but no absolute conclusions, about a great number of alkaloids so far known of this group, and facilitate the identification of some of the more common bases. 3. No absolute relation between RM value and structure can so far be derived from the RC values of Strychnos alkaloids. This is due to the extremely complicated structures of these alkaloids, which belong to several chemical groups, and to the difficulty of obtaining standard RC values for many quaternary alkaloids. 4. Paper electrophoresis of Strychnos tertiary alkaloids is of great importance for the identification of the different strychnine derivatives. 5. Paper chromatography and electrophoresis must be considered as the best methods hitherto used for the preparative purification of small quantities of Strychnos quaternary alkaloids.

A silicic acid paper chromatographic method for the separation of four major classes of cholesterol esters, namely saturated cholesterol esters, cholesterol oleate, cholesterol linoleate and cholesterol linolenate plus cholesterol arachidonate is described. This method uses a benzene-hexane solvent. The use of combined column and paper chromatography as well as gas chromatographic analysis is presented for definitive identification of the various spots. The predominance of cholesterol linoleate in human plasma, and cholesterol arachidonate in rat plasma is confirmed.

1.1. Filter paper electrophoretic analysis of the serum and urine proteins in thirty-five cases of multiple myeloma has been carried out. In all thirty-five cases characteristic protein abnormalities were observed; seventeen cases showing homogeneous peaks in both the serum and urine, eleven cases in serum only, seven cases in urine only. 2.2. All urine samples showing a positive Bence-Jones protein reaction also displayed a characteristic electrophoretic abnormality. Eight of the twenty-four electrophoretically positive urine samples were Bence-Jones protein negative by the usual tests. Because of the relative simplicity of urine electrophoresis by the filter paper technic, this test is considered to be a practical clinical procedure. 3.3. Two cases are presented in detail because certain atypical clinicopathologic features of their disease patterns possibly assist in an understanding of the relationship of the plasma cell dyscrasias to chronic lymphatic leukemia and the lymphosarcomas. A third case, which demonstrates the value of serial electrophoretic studies in myeloma, is also presented. 4.4. Consideration is given to some of the physicochemical properties of the myeloma serum and urine proteins, and to the possible nature of their interrelationship.

A paper Chromatographic method for the analysis of phenylalanine in serum is described. The amino acids, including phenylalanine, are isolated from the deproteinised serum by means of a cation exchanger. The phenylalanine is then separated by paper chromatography at elevated temperature and estimated colorimetrically. Normal values are given. The excretion of phenylalanine and phenylpyruvic acid is correlated with the serum phenylalanine level. The method is specific and the reproducibility is good. Its application in a case of phenylketonuria is reported.

Inorganic anions were studied by paper electrophoresis and ion-exchange paper chromatography employing electrolytes and eluents with various metal ions (Mg2+, Al3+ and Cd2+). It was found that the metal ion has a strong effect on the movement of anions in paper electrophoresis both by ion pair and complex formatin In ion-exchange paper chromatography the effect is small. Data are given for twenty inorganic anions.

The behaviour is described of several barbiturates when subjected to chromatography on filter paper impregnated with olive oil. Results are given for various methanolic and aqueous inorganic solvents. The technique affords a fairly rapid separation of slow-, medium- and quick-acting barbiturates but the order of separation is different from those obtained with conventional and ion-exchange paper chromatography.

〓 (R=2-サフトール-6, 8-ジスルホン酸, 2-ナフトール-3, 6-ジスルホン酸, 2-ナフトール-6-スルホン酸, 2-ナフトール-7-スルホン酸, 2-ナフトール-8-スルホン酸) についてスルホン酸基の置換位置または数と塩析効果の受けやすさとの関係を, 塩化カリウム水溶液または含アルコール, 塩水溶液を展開液とした塩析型ロ紙クロマトグラフィーのRf値等により検討した。さらにベンザルレブリン酸の3種の位置異性体をその2, 4-ジニトロフェニルヒドラゾンにしたものをロ紙電気泳動または塩析型ロ紙クロマトグラフィーにより明確に分離した。

Paper-electrophoretic separation of intrinsic factor-related “primary” and “secondary” B12 binders in pooled normal human gastric juice was achieved by using Teorell-Stenhagen buffers of pH 4.0–5.0, at which pH the “primary” binder moves to the cathode, while the “secondary” one travels to the anode. The isoelectric point of the “primary” binder is about pH 5.5, while that of the “secondary” one is pH 2.5–3.0. The electrophoretic mobility of none of the major carbohydrate-, proteinor peptide-nondialyzable components of gastric juice correlates with that of the “primary” or “secondary” B12 binders when run on electrophoresis in the pH range of from 2.5 to 10.0. This supports previous conclusions that none of the major components of gastric juice resolved by paper electrophoresis represents the intrinsic factor-related B12 binders.

Assumptions concerning the similarity of paper chromatography and paper electrophoresis as regards the nature of the interaction of the substances analysed with the immobile phase were verified. glueckauf's method for constructing distribution isotherms from chromatographic measurements was applied to electrophoretic measurements. It was proved that the isotherms obtained in this way are identical with the isotherms obtained by the direct method, when the dependence of the mobility in free solution on the concentration can be neglected. The values of RF, the electrophoretic mobility in free solution Uo and the velocity uo obtained both by direct measurement and by calculation were proved to be identical.

The possible degradation of tryptophan and seven of its metabolites during and after paper chromatography with n-butanol-acetic acid-water has been investigated. It has been found that extensive degradation results from developing and drying the chromatograms at temperatures over 27° and storing them in room light for extended periods before the second development. Lesser amounts of degradation were found after limited exposure of compounds to ultraviolet light between developments and occasionally with the use of unwashed paper. Procedures to eliminate degradation have been described.

A study has been made of some of the fundamental factors involved in obtaining good separations and reproducible chromatograms, with the technique of contrifugally accelerated paper chromatography. A study was also made of variables which might influence R F values. Mixtures of bromphenol blue, methyl orange and methyl red were fractionated as well as solutions of lencine, methionine and glycine. For the separation of the dyes, Whatman No. 1 filter paper was used with a veronal buffer solution of ionic strength 0.05 and having a pH of 8.7. For this system, it was found that the R F values of the dyes were essentially unaffected by changes in rotational speed of the rotor from 300 to 925 r.p.m. It was found that the rate of addition of solvent was important in developing satisfactory chromatograms. A flow rate of 1.15 ml/min was found to be satisfactory at all rotational speeds above 250 r.p.m. The shape of the developed chromatogram was elliptical rather than circular, for all four types of paper which were studied. For Whatman No.1, the major axis of the ellipse was parallel to the machine direction of the paper. The R F values for the dye components were essentially the same for the following papers: Whatman No. 1, Eaton-Dikeman No. 613, and Cremer-Tiselius-Munktells. The amino-acid mixture was fractionated using Whatman-3MM paper and a solvent system of butanol-glacial acetic acid and water, in the volume ratio 40:10:10, respectively. The disk was rotated at 975 r.p.m. With a solvent flow rate of 1.2 to 2.1 ml/min, sharply defined zones were obtained. The time of development was 15 min for a flow rate of 1.2 ml/min and 7 min for a flow rate of 2.1 ml/min. The R F values obtained were 0.85 for lencine, 0.56 for methionine and 0.20 for glycine.

On account of differences in lipophilic character it is possible to separate p -nitrophenyl methylphosphonates and their sulfur analogs by reversed phase paper chromatography. Whatman No. 1 filter paper was impregnated with silicone oil 550; the upper layer of a chloroform-ethanol-water (5:5:3, v/v) system served as mobile phase. The relation R M = log (1/ R F —1) holds for homologous series of O-alkyl p -nitrophenyl methylphosphonates (I) and S-alkyl p -nitrophenyl methylphosphonothiolates (II) up to the n -pentyl homologues. Using a combination of Gibbs' and Dragendorff reagents it was possible to distinguish the phosphonates, phosphonothiolates and phosphonothionates.

A study with filter paper strips partially impregnated with the precipitating agent with or without colloid as the fixing agent reveals that in the case of spotted strips run in aqueous alcoholic mixture as the solvent, uniform bands of the precipitate are formed. In these cases, chromatographic observations such as measurement of ‘S’ and ‘Rf’ values is possible. Rhythmic structures can also be obtained with these strips when the reactant solution is used as the solvent. In the absence of colloids rhythmic structures are obtained at a high concentration and the bands obtained have to be washed and repeatedly dipped in hot water in order to clear the structure of rings present underneath. In the presence of colloid, rhythmic vertical bands are obtained at a much lower concentration and the clear interspaces are visible even without washing. Thus an independent method for obtaining rhythmic structures on filter paper with a technique very similar to chromatography has been developed.

After conversion to 2,4-dinitrophenylhydrazones, keto acids in biological fluids were extracted into isopropyl ether followed by NaHCO 3 and finally into ethyl acetate. The derivatives were chromatographed two-dimensionally on Whatman No. SG 81 paper. Limitations of the extraction and chromatographic procedures are discussed but it is considered that the method is superior to orthodox one-dimensional paper chromatography.

Following previous supplements in this series, a further 180 compounds were analyzed by paper chromatography in six solvent systems. The data originating from 15 standard colour reagents are also recorded and presented. The following types of compounds are described: flavonoid derivatives, aliphatic and aromatic aldoximes and ketoximes, substituted benzamide derivatives, aminobenzene derivatives and heterocyclic amines, N-nitroso compounds, N-oxides of tertiary bases, pyrimidine and indole derivatives and sulphonamide drugs. The paper chromatographic mobility patterns were characteristic, especially for 1,3- and 1,4-hydroxybenzaldoximes and ketoximes of the hydroxyacetophenone type. All the heterocyclic bases investigated showed mobility patterns similar to those of the alkaloids described in earlier supplements. Mobility patterns similar to the alkaloids were produced by some of the mercaptopyrimidines. In general amino-, diamino-, hydroxy and dihydroxy derivatives of pyrimidines showed low mobility in these solvents. Several flavonoid compounds were found to be Erhlich positive and a magenta colour was produced by l -epicatechin. Most of the N-unsubstituted benzamides were detected with p -dimethylaminocinnamaldehyde, and most of the N-nitroso derivatives of secondary amines and N-oxides of heterocyclic tertiary amines were revealed by this reagent.

A paper chromatographic solvent system and an extraction and concentration microtechique were used to determine air-borne 1,2:3,4-dibenzopyrene, 3,4-benzofluoranthene, 3,4-benzopyrene, 1,12-benzoperylene and anthanthrene in an automobile tunnel, in frequented cross roads, ad in sites with little or no automobile traffic. Fluoranthene, pyrene, chrysene, 1,2-benzanthracene, 1,2:5,6-dibenzanthracene, 11,12-benzofluoranthene and coronene were identified on the same chromatogram, 1,2-benzopyrene was presumed. Using at least three filters in series, quantitative air-sampling of penta- and higher cyclic hydrocarbons is claimed. The results are tabulated.

A two-dimensional paper chromatographic method is described which allows good separations and spectrophotometric determinations of nanomole amounts of hypoxanthine, xanthine, uric acid and uric acid riboside. The method can be applied to biological material containing adenine nucleotides and considerable amounts of free amino acids.

The RF values of inorganic ions on paper pre-treated with 0.25–2.0 % cellulose acetate and impregnated with various concentrations of 2-thenoyltrifluoroacetone (TTA) was studied by using acetate buffer as a developing agent. 1% cellulose acetate pre-treated paper impregnated with o.2 M TTA was found suitable for reversed phase paper chromatography. The RF values of inorganic ions were examined on the impregnated paper by using various pH's of the developing agent (0.1 M acetate buffer). Separations of PbCu, CoCu, ZnCu and FeCoNi were performed in this system.

The results obtained with freshly prepared solutions indicate that ferric ions may be adsorbed by Whatman No. 1 filter paper by a process of ion exchange. However, on standing the system becomes complicated by the diverse nature of the hydrolysis products formed. Increased adsorption takes place, apparently through the retention of hydroxy compounds of lower net charge.

The frontal method of measuring chromatographic distribution isotherms was applied to paper electrophoresis. Measurements were performed for a model coloured substance. Concentration dependence of free electrophoretic mobility, of electrophoretic mobility on paper, and of RF were obtained.

The mammalian respiratory tract produces a secretion in which the major macromolecular components are mucin-type glycoproteins. High-voltage paper electrophoresis has proved a valuable research tool in studies of the formation of these glycoproteins. Separations of hexoses, amino sugars, oligosaccharides, sugar nucleotides and mucin glycoproteins are rapidly and easily obtained by paper electrophoresis at voltages of 50 to 100 V/cm. The use of high-voltage paper electrophoresis to follow amino sugars interconversions and the transfer of glycosyl units to mucin acceptors is described.

A rapid Kunkel type electrophoretic system was used to determine the effect of wetness and temperature upon the mobility of serum components. Comparison is made with a Durrum type apparatus. Variability averaged 6.5% for albumin with both techniques. Observed component mobility increases both with wetness and temperature with the Kunkel system. When corrected by use of dextran, observed mobility values are independent of wetness, but increase directly with temperature. Comment is made on mobility markers and a wetness ratio, on specimen identification by mobility alone and on the usefulness of zone electrophoresis in qualitative analysis.