Clx. a Study of Seasonal Variation in Butter-fat. I. Seasonal Variations in Carotene, Vitamin a and the Antimony Trichloride Reaction. by Roland

IN January 1932 work was started in the National Institute for Research in Dairying to investigate the variations in the vitamin content of typical English milk as determined by the season of the year and the nutritional condition of the cow. This work, which was planned to cover two complete years, and which will be described in detail later, is mainly concerned with the accumulation of data obtained in routine biological tests carried out according to accepted methods. In several instances however unexpected difficulties have been encountered, and it has been necessary to seek new explanations of the observed facts. Among other occasions this was the case when it was attempted to apply the well-known colorimetric and spectroscopic methods to the estimation of carotene and vitamin A in butter-fats, and the present joint work with the Nutritional Laboratory is concerned with our attempts to overcome this difficulty. In assaying the vitamin A of cod-liver oils by the antimony trichloride method it is well known that the blue value calculated per unit of the original oil may be greatly increased as the result of saponification. Green [1932] and Moore [1932] have already noticed that the blue value given by butter-fat is similarly increased on saponification, and this finding has been confirmed in the National Institute for Research in Dairying in routine tests on over 150 samples of butter-fat. What specially attracted our interest however was the fact that the blue value was not always increased in the same ratio, and that in general the increase was much greater in samples of butter obtained from cows at pasture ("summer butters") than in samples obtained from stall-fed cows receiving diets totally or partially deficient in green fodder ("winter butters"). In addition it was noticed that the colours produced by the untreated winter and summer butter-fats differed qualitatively. The maximum intensity of blue in the case of winter butter was given immediately on the addition of the antimony trichloride reagent, after which fading took place very rapidly (2-5 secs.) without the formation of any intermediate tint. With summer butters on the other hand the blue colour was usually more stable, sometimes taking a few seconds to reach its maximum value, while fading was preceded by the change in the colour of the solution from the original greenish-blue to a dull reddish-blue. Now it is well known that the blue colour given by vitamin A in the antimony trichloride test is "inhibited," a term which may be considered to embrace diminution in intensity, alteration in tint or acceleration in the