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THE formation of more than one zone containing phosphate when the sodium orthophosphates are chromatographed on paper suggests that these spots contain different molecular species originating in proton transfer equilibria of the kind: where B−nis a base1,2. The speed of proton transfers, for example, the half-life for the reaction of hydronium and hydroxyl ions is 10−8sec3, argues against this as such equilibria yield multiple spots only if the transition time is comparable with the time required for the formation of the chromatogram4. It is natural to question next if the zones are actually of different species since discontinuities in either the immobile or mobile phases can also produce the result4. Chromatograms of orthophosphoric acid and of the primary, secondary, and tertiary sodium salts were formed on 55 cm × 2.8 cm strips of Whatman No. 1 filter paper, acid washed, which had also been washed in 2 M acetic acid followed by three successive rinsings in distilled water5. The chromatograms were formed either with pyridine/ethyl acetate/water (the faster solvent) or butanol/water. Fig. 1 is a photograph of 10 µl. of 0.01 M trisodium phosphate (No. 321) and 10 µl. of 0.01 M sodium dihydrogen phosphate (No. 322) formed simultaneously with BuOH/15 per cent H2O. The origin was the vertical pencil line adjacent to the numbers. When revealed, the colour process was that employing ammonium molybdate and benzidine hydrochloride6. In certain select cases, the multiple spots were irradiated in the University's Triga reactor7,8. Both sodium and phosphorus undergo (n,γ)·reactions to form radioactive isotopes which decay as follows9,10:

THE applications and limitations of one- and two-dimensional paper chromatographic techniques have been well documented1. There remain, however, problems in the separation of certain compounds which cannot bo resolved satisfactorily by either of these conventional approaches. Weiss and Rossi2recently described a chromatographic method which enables separation of several catecholamines on a single paper strip.

IN translating this paper of Maxwell for Ostwald's “Klassiker der exacten Wissenschaften,” I have detected some errors, which are partly merely misprints, but partly also faults in the formulæ of some trouble to the reader. The German translation is only of value to those who have not leisure to study the English language before the works of Maxwell; but the accuracy of such classic works is so essential to every one, that I considered the publication of the errata found to be of even greater importance than my whole translation. But in order to make the translation as cheap as possible, the German editor refused to print my list of errata, and I therefore hope it will be printed in England.

TEMPERATURE is a variable which has received relatively little attention in chromatographic studies. This is particularly true of paper chromatography in which more attention is given to controlling temperature as an aid to reproducibility than to using temperature to effect improved resolution.

LEIFER et al.1have reported the paper chromatography of nicotinic acid with the use of microbiological assay. While our work was in progress, Huebner2described a spot test for detection of some pyridine derivatives. The present work describes sensitive qualitative techniques developed in this Laboratory for the detection of tertiary and quaternary pyridinium compounds. The laborious procedure involved in the microbiological method of Leifer et al.1is obviated by rapid colour or fluorescent tests. For the colour test, advantage was taken of König's3reaction, namely, the formation of coloured compounds when tertiary pyridine derivatives are treated with cyanogen bromide and a primary aromatic amine. The fluorescence test allows the detection of quaternary nicotinamide derivatives. Use is made of the formation of bluish-white fluorescent substances when quaternary pyridine compounds with a side chain CO—NH—R in the β position are treated with methyl ethyl ketone and ammonia4.

WE have shown1that primycin, an agent of strong antituberculotic effect belonging to the group of guanidino antibiotics, is not identical with any of the known antibiotics. On the basis of its colour reactions and of its infra-red and ultra-violet spectra2, the molecule of primycin (C19H37O7N)ncontains an N—H group and several O—H groups; it is inclined to molecular associations and it discloses adsorptive properties. On subjecting primycin to paper chromatography, the formation of a second spot was observed. This was due to the so-called ‘secondary adsorption’ of an accompanying substance present in minute amounts.

I. —A VERY COSTLY AND VEXATIOUS FALLACY

CHROMIUM sesquioxide (Cr2O3) is widely used for estimating feed digestibility1or fæces output2by ruminants, but such estimates may be subject to considerable errors because the concentration of this marker in fæces normally shows considerable diurnal variations even after prolonged regular administration. These variations are particularly obtrusive with grazing animals where the chromium sesquioxide, in this case used for estimating the output of faeces, is usually administered in concentrated powder form in gelatin capsules once or twice daily, and they probably arise primarily from an imperfect mixing of the oxide with digesta in the reticulo-rumen3,4. This view is supported by unpublished observations (by J. F. D. Greenhalgh) on the concentrations of chromium sesquioxide in the organic matter of digesta obtained from the duodenum of a sheep permanently cannulated at this point. As the variations in concentration during a period of regular dosing showed the same pattern and were greater than those found in the fæces, it was decided to study in a comparably cannulated sheep the passage of single doses of oxide powder, each given in a gelatin capsule and in amount large enough to give measurable concentrations over a considerable period of time. Samples of digesta were drawn from the duodenum at intervals beginning 1 hr. after dosing. Results from a typical run where 8 gm. were given are shown in Fig. 1 (line A) and indicate that a large part of the dose had passed from the stomachs within 4 hr. of administration. The probable explanation is that the capsule container dissolved quickly in the anterior rumen or reticulum3and that local concentrations of chromium sesquioxide in these regions4were rapidly transferred through the reticulo-omasal orifice.

METABOLITES of numerous biologically active endogenous and exogenous organic compounds are excreted in bile and urine as glucuronic acid conjugates. In the course of studying the mechanism of action of carcinogenic polycyclic hydrocarbons, we wished to identify the metabolites of certain of these agents. For this purpose, 14C-3,4-benzpyrene was injected into rats, and bile was collected by an indwelling cannula, utilizing a technique already described1. The bile fractions were collected in an automatic fraction collector, and enzymatic hydrolysis with β-glucuronidase was undertaken. Characterization of all the metabolites by this technique proved to be impossible because of the extreme instability of some of the hydrolysis products. We therefore attempted to determine whether the extent of oxidation and the class of conjugate formed could be established by direct analysis of the conjugates.