IL D. Trw~I and K. C. SRIVASTAVA: Paper Chromatography of Fatty Acids 117

ten rostfreien Stahlrohr selbst bei Wasserstoffgehalten yon fiber 12~ Leitungsl/~ngen bis zu mehreren Metern ohne Beheizung gestattet und so die Nullpunktsstabilit/~t der Waage nicht gef~hrdet. Die l~esultate mit diesem Ger~t sind sehr erfolgversprechend und es wird an der technischen Vervollkomnmung gearbeitet.

Literatur 1. T~vT~OVSKY, H. : diese Z. 22~, 254 (1966). 2. -- Mikroehim. Acta (ira Druek).

Dr. H. TI~UTNOVSKu Med.-chem. Institut u. lh'egl-Laboratorium Graz, Universit/~tsp]atz 2 (0sterreieh)

Paper Chromatography of Fatty Acids Use of Acetates of Cobalt, Copper and Nickel and of Mercurous Nitrate as Spot Detect ing Reagents

R. D. TIWA~I and K. C. S~IVASTAVA

Chemistry Department, University of Allahabad, Allahabad (India)

Received March 1, ].967

In the present communication separation of saturated fa t ty acids (C1~ to C2o ) has been achieved by reversed-phase partition chromatography using the descending technique. The impregnating materials used are li- quid paraffin (10--12~ w/v in benzene) and a benzene solution of 0.5~ of cetyl alcohol. The use of the second impregnating material makes the first one a good adsorbent with the only reservation that its quanti ty should be kept below that. The developing solvent used is a mixture of acetone and methanol (1:3 by volume). The acids have been revealed by treating the developed paper with aqueous solutions of (a) cobalt acetate [1] followed by treatment with either an aqueous solution of nitroso-R- salt or an aqueous solution of sodium diethyldithiocarbamidate; (b) cop- per acetate followed by treatment with either an aqueous solution of sodium diethyldithiocarbamidate or with water saturated with hydrogen sulphide gas; (c) nickel acetate followed by treatment with either an aqueous solution of potassium hexacyanoferrate(II) or an alcoholic sohltion of dimethylglyoxime and (d) mercurous nitrate followed by treatment with an aqueous solution of gallic acid.

Experimental Procedure. Whatman No. 1 chromatographic paper of a suitable size was impregnated by dipping it into a solution of liquid paraffin and cetyl alcohol in bezene (12~ and 0.3--0.5 ~ respectively). It was removed from the solution and hung vertically whereupon the solvent evaporated at room temperature. Spots of the fatty acids

118 R.D. TzWARZ and K. C. SRZVASTAVA: Paper Chromatography of Fatty Acids

as their benzenic solutions were applied on the paper Mong with that of their mixture with the help of a mieropipette. The paper was then developed with a mixture of acetone and methanol (1:3 v/v) using the descending technique. After the solvent front had moved a requisite distance, the paper was dried with a hair drier and was further subjected to the following treatments. a) I t was placed in an aqueous solution (4--5~ of cobalt acetate [1] for about 10-- 15 rain. I t was removed from this solution, washed thoroughly with tap water and then placed in another tray and sprayed with 0.10/0 aqueous solution of either nitroso-R-sMt or sodium diethyldithiocarbamidatc. When sprayed with the first reagent the acids appeared as orange-red spots with light orange background on the entire paper. As the complex formed between cobalt and nitroso-l%-salt is a water soluble one, much of the undesirable colour of the back-ground could be removed by gently washing the paper with tap water. A second spray with the same reagent, however, revealed the acids as orange-red spots against a faint orange back-ground. When sprayed with the second reagent (sodium diethyldithiocarbamidate) the acids were located as dirty green spots against a very light green back-ground of the paper. This time a rewashing followed by another spray with this reagent did not, however, improve the intensity of the spots probably because the complex formed between bivalent cobalt and this reagent is not soluble in water. b) The paper was placed in an aqueous solution (5--7~ of copper acetate for about 15 rain, washed thoroughly with tap water and then either sprayed with an aqueous solution of sodium diethyldithiocarbamidate or dipped in water satu- rated with hydrogen sulphide gas. With the first reagent the acids appeared as black-brown spots while with the second one they appeared as black spots. c) The paper was placed in an aqueous solution of nickel acetate (7--8~ for about 15 rain, washed thoroughly with tap water and then dipped into an aqueous solution (10~ of potassium hexaeyanoferrate(II) for 5 rain. The acids appeared

as green-blue spots. A better way of revealing the Table acids was to spray the nickel acetate treated paper Rf-values o//atty acids washed thoroughly with tap water, with an l~ Acids Rt-values alcoholic solution of dimethylglyoxime followed by a

spray with 50 ~ ammonia in water. The red complex Lauric 0.994 between nickel and dimethylglyoxime intensifies in Myristic 0.902 the presence of ammonia. The entire paper also Palmitic 0.821 turned red but the intensity of the colour was much Stearic 0.683 higher at the site of the spots. To get rid of the Arachidic 0.624 back-ground eolour it is desirable to dip the paper

into a 30~ aqueous solution of glacial acetic acid for 5 rain or less. On such a treatment the colour of the spots dimmed with the disappearance of the back-ground colour giving a better identity of the acids than when the back-ground colour was present. d) The paper was treated with an aqueous solution of mercurous nitrate for 10 to 15 rain, washed thoroughly with water and then sprayed with 0.50/0 aqueous solution of gallic acid. The acids were located as orange-yellow spots.

Rf-Values of different f a t t y acids are shown in the Table.

Reference

1. Tr~v~i, R. D., and K. C. S~rVAS~VA: Z. Anal. Chem., 230, 36 (1967)

Dr. K. C. SRIVASTAVA Department of Chemistry, University of Allahabad Allahabad (India)