Study on the interaction of proteins and carbohydrates and its bearing on the nurtitive value

Abstract

Synopsis of the thesis entitled “Studies on the Interaction of Proteins and Carbohydrates and Its Bearing on the Nutritive Value” submitted by Mr. P. B. Bamino, B.Sc. (Hons), for the degree of Doctor of Philosophy of the University of Bombay. The spontaneous deterioration of stored skim milk powder reported by Henry, Kon, Lea and White (1945) has opened an entirely new field of research concerning proteins and carbohydrates. The deterioration of stored foodstuffs was normally attributed to causes such as high moisture content, exposure to air and light, and bacterial contamination. Adequate precautions taken against these known factors during processing and subsequent storage were considered sufficient for the proper preservation of foods. However, the protein–sugar reaction taking place in the “dry” state, resulting in the deterioration of protein quality, is a new factor to be tackled by food technologists. A systematic study of the exact conditions under which it is brought about and an assessment of its nutritive effect will be of great help. Maillard showed that amino acids and reducing sugars reacted. This observation related to free amino acids and sugar was perhaps not believed to occur with proteins—until Henry, Kon and co?workers published their results on the spoilage of stored milk powder. Following this report, many investigators also published their results, and these studies have greatly improved our understanding of the problem. Though connected mainly with the deteriorative aspect, the reaction also presents certain desirable features. The formation of the brown crust colour in bread is a result of this reaction, and in certain cases when the desirable crust colour was not formed, addition of egg protein facilitated the rapid development of the crust colour. Many cooked or processed foods such as toasted cereals, toffee, roasted coffee, malted barley, etc., owe their aroma, flavour and colour to the result of this reaction. Further, studies with free amino acids and sugars have shown the possibilities of producing new or modified flavours. But the deteriorative aspect of the reaction is more serious than the possible advantages, for the nutritive value of the protein is so much affected as not to support life. Lea and Hannan (1950) conducted systematic studies to evaluate the conditions of temperature, hydrogen?ion concentration, humidity and the type of reducing sugar that aided the reaction. Henry and Kon (1950) observed that casein–glucose mixtures stored for 30 days had decreased in biological value and digestibility to an extent at which the protein could hardly be considered adequate for nutrition. Patton and co?workers (1948) reported the loss of a variety of essential amino acids to varying extents when proteins were heated in solution or autoclaved with reducing sugars. The lowering in the availability of the amino acids to micro?organisms was traced to the actual loss of the amino acids. Reader, Stewart and Smith (1949) observed that it was not so much the loss of amino acids that was responsible for lowering the nutritive value of protein as the reduction in digestibility. Krafft and Morgan (1951) and Harvey and Guggenheim (1953) reported that the nutritive value of the protein–reducing?sugar complex was restored to its original level by supplementation with lysine. The present author carried out a systematic study of the entire problem, and his observations are recorded and discussed in this thesis. The author successfully applied paper chromatography to study the extent to which amino acids were affected as a result of the reaction with reducing sugars. The reliability of the microbiological assay of amino acids in hydrolysed protein–sugar complexes has been questioned by previous workers, as large discrepancies were pointed out regarding availability of the amino acids to micro?organisms compared to their availability to rats from the same protein–sugar complex. The successful application of paper chromatography is therefore shown to be particularly advantageous. The extent to which different amino acids were lost when proteins were autoclaved with reducing sugars showed that only the loss of lysine, of the essential amino acids, was serious. Arginine, a non?essential amino acid, was shown to be completely destroyed under the conditions employed, and the absence of the arginine band from the chromatogram was conspicuous. The other basic amino acid, histidine, was also affected. Studies on the degree of loss in increasing time of autoclaving showed that the maximum loss occurred with 4 hours of autoclaving, and further autoclaving of the protein–glucose mixture did not result in any further significant loss of any of the amino acids. Biological value determinations by the balance?sheet and protein?efficiency?ratio methods indicated that both the protein value and digestibility were affected. Supplementation with lysine restored the nutritive value of the protein–glucose complex to the original level. These observations were substantiated by in?vitro studies. These experiments, carried out by the author for the first time, showed that the rate of digestion in vitro of the protein–glucose complex by trypsin was improved by the addition of lysine. It may be stated that in the reaction between the protein and the reducing sugar, the terminal amino group is blocked, as a consequence of which the enzyme does not gain access to the protein. Addition of lysine possibly gives rise to the re?formation of the active group in the protein molecule, necessary for proteolytic activity, or it may counteract the inhibitory influence of the products of heat degradation of the sugars or the reaction products on trypsin activity, improving the digestion rate. Experiments were also conducted to confirm the usefulness of lysine supplementation by other methods of evaluating protein quality. The capacity of the protein–glucose complex for haemoglobin regeneration and liver?protein formation—with and without supplemental lysine—gave further proof that the resistance of the protein–sugar complex to enzymic digestion, which could be overcome by lysine supplementation, together with the loss of the amino acids themselves, was responsible for the deterioration of protein quality. The deteriorative aspect of the reaction was seen less in the case of intact cereals and pulses. It was felt that starch could exert a protective action on the protein. Experiments were therefore carried out with casein to which different amounts of starch were added. Autoclaving with glucose of the mixture showed that the extent of loss of amino acids decreased with increasing percentage of starch in the mixture. The effect of spray?drying milk and processing milk?sugar products by indigenous methods was also studied to determine the amino?acid losses.