Casein

Abstract: The article summarizes research into the existing methods for the quantitative determination of tyrosine and tryptophane in proteins. Limitations to the accuracy of the Folin-Looney method (reaction of a phosphotungstic phosphomolybdic acid in a phenol solution, evaluated using colorimetry) have been solved by an improved method detailed in the text. The hydrolysis of proteinaceous material to allow chemical analysis of tryptophane has also been improved; the method is based on digestion with sodium hydroxide for 18-20 hours, followed by rapid neutralization and acidification with sulfuric acid. A more accurate test for tyrosine based on Millon's reaction has been developed; acidified mercuric sulfate solution is used to dissolve precipitated tyrosine and sodium nitrite is added to produce the colored product which is assessed colorimetrically. Two types of casein analyzed by these methods contained 1.4% tryptophane and 6.37-6.55% tyrosine. Tryptophane and tyrosine content of various materials were: casein 1.4%, 6.4-6.6%; egg albumin 1.3%, 4.0%; edestin 1.5%, 4.5%; gliadin 0.84%, 3.1%; zein 0.17%, 5.9%. A method for preparation of the pure mercuric sulfate reagent is described.

Abstract: Catalyzed phenol-hypochlorite and ninhydrin colorimetric procedures were adapted to the Technicon AutoAnalyzer for simultaneous determination of ammonia and total amino acids in ruminal fluid or ruminal in vitro media. The manifold developed was compatible with a sampling rate of 40/h without significant sample-to-sample carryover. With proper storage, reagents for both the phenol-hypochlorite and the air-stable ninhydrin systems were stable for 8 mo or more. Response of individual amino acids in the phenol-hypochlorite system were generally 1% or less than equimolar amounts of ammonia. Certain amino acids inhibited ammonia color yield 10 to 15% when with equimolar amounts of ammonia; however, the inhibitory effect of casein amino acids was only 2 to 3%. Although ninhydrin response, relative to leucine, of individual alpha-amino acids ranged from 62 (valine) to 151% (histidine), recoveries of casein amino acids from ruminal fluid had coefficients of variation of 1% or less. Coefficients of variation for ammonia recoveries from ruminal fluid by the phenol-hypochlorite procedure were about half of those for the Conway microdiffusion technique. Intraclass correlations for the adapted procedures indicated high degrees of accuracy and precision for both ammonia and amino acid analyses.

Abstract: There has been a great need for a satisfactory method for the determination of hydroxyproline to facilitate the study of the composition of proteins. Lang (1) and Waldschmidt-Leitz and Akabori (2) developed a calorimetric estimation involving oxidation to pyrrole with sodium hypochlorite and color formation with isatin or p-dimethylaminobenzaldehyde. However, oxidation was said to be incomplete and a correction factor was necessary. Dakin (3) and Bergmann (4) estimated hydroxyproline by isolation procedures. These procedures also required the use of large correction factors, as well as relatively large quantities of protein. McFarlane and Guest (5) devised a calorimetric method for hydroxyproline involving sodium peroxide oxidation and color formation with copper and isatin. This method yielded low values according to Devine (6), and in our hands similar results were obtained. The unique high hydroxyproline content of collagen suggests the desirability of an accurate method for the determination of this amino acid in small quantities as a means of estimating the amount of collagen or gelatin in a mixture of proteins. Because hydroxyproline so far has not been found to be a nutritive requirement for a microorganism, a chemical procedure is required. A short and simple calorimetric method is here described that is applicable to the determination of hydroxyproline in hydrolysates of 40 to 100 y of collagen with a reproducibility of f2 per cent (Table II) and an accuracy of f2 per cent (Table I) as judged by recovery of hydroxyproline from elastin hydrolysates and from an amino acid mixture simulating collagen. Oxidation, in the manner of McFarlane and Guest with sodium peroxide, yields products that form an intense red color with p-dimethylaminobenzaldehyde. The intensity of color produced with 5 to 15 y of hydroxyproline is 4 to 5 times that formed in previous colorimetric procedures. Because of this greater intensity of color and because of a different preliminary treatment of the hydroxyproline solutions, only 1 to 2 per cent as much protein is required as in earlier methods. In acid hydrolysates of proteins, the only amino acid other than hy-

Abstract: Physico-chemical characteristics of milk are related to its composition for a particular animal species. Sheep milk contains higher levels of total solids and major nutrient than goat and cow milk. Lipids in sheep and goat milk have higher physical characteristics than in cow milk, but physico-chemical indices (i.e., saponification, Reichert Meissl and Polenske values) vary between different reports. Micelle structures in goat and sheep milk differ in average diameter, hydration, and mineralization from those of cow milk. Caprine casein micelles contain more calcium and inorganic phosphorus, are less solvated, less heat stable, and lose β-casein more readily than bovine casein micelles. Renneting parameters in cheese making of sheep milk are affected by physico-chemical properties, including pH, larger casein micelle, more calcium per casein weight, and other mineral contents in milk, which cause differences in coagulation time, coagulation rate, curd firmness, and amount of rennet needed. Renneting time for goat milk is shorter than for cow milk, and the weak consistency of the gel is beneficial for human digestion but decreases its cheese yield. Triacylglycerols (TAG) constitute the biggest part of milk lipids (nearly 98%), including a large number of esterified fatty acids. Sheep and goat milk also have simple lipids (diacylglycerols, monoacylglycerols, cholesterol esters), complex lipids (phospholipids), and liposoluble compounds (sterols, cholesterol esters, hydrocarbons). The average fat globule size is smallest ( 75% of total fatty acids in goat and sheep milk. Levels of the metabolically valuable short and medium chain fatty acids, caproic (C6:0) (2.9%, 2.4%, 1.6%), caprylic (C8:0) (2.6%, 2.7%, 1.3%), capric (C10:0) (7.8%, 10.0%, 3.0%), and lauric (C12:0) (4.4%, 5.0%, 3.1%) are significantly higher in sheep and goat than in cow milk, respectively. Principal caseins (CN) in goat, sheep and cow milk are αs1-CN, αs2-CN, β-CN and κ-CN. The main forms of caprine and ovine caseino-macropeptides (CMP), which are the soluble C-terminal derivatives from the action of chymosin on κ-casein during the milk clotting process of cheesemaking, have been identified and are a good source of antithrombotic peptides. Sheep and goat milk proteins are also important sources of bioactive angiotensin converting enzyme (ACE) inhibitory peptides and antihypertensive peptides. They can provide a non-immune disease defence and control of microbial infections. Important minor milk proteins include immunoglobulins, lactoferrin, transferrin, ferritin, proteose peptone, calmodulin (calcium binding protein), prolactin, and folate-binding protein. Non-protein nitrogen (NPN) contents of goat and human milks are higher than in cow milk. Taurine in goat and sheep milk derived from sulphur-containing amino acids has important metabolic functions as does carnitine, which is a valuable nutrient for the human neonate. Mineral and vitamin contents of goat and sheep milk are mostly higher than in cow milk.

Abstract: This report reviews changes the nomenclature of bovine milk proteins necessitated by recent advances of our knowledge. Identification of a number of milk proteins ( α s1 -, β -, and κ -caseins; α -lactalbumin and β -lactoglobulin) continues to be based upon their primary structures (amino acid sequences). Since our last report, α s2 -casein and serum albumin can be added to the list of major milk proteins for primary structure is known. Changes recommended in the nomenclature of caseins are primarily a result of differences within this family of proteins brought about by posttranslational modification. For example, α s0 -casein is identical to α s1 -casein, and α s3 -, α s4 -, and α s6 -caseins are identical to α s2 -casein except for differences in degree of phosphorylation. Additionally, proteose-peptone components 5, 8-slow and 8-fast, and γ 1 -, γ 2 -, and γ 3 -caseins are N-terminal and C-terminal fragments, respectively, of β -casein formed during proteolysis by plasmin. Nomenclature of immunoglobulins remains consistent with guidelines for human proteins and is based largely upon crossreactivity with reference proteins. The minor whey protein lactollin is β 2 -microglobulin for which the sequence of amino acids is known. An operational definition for proteins associated with the milk fat globule membrane has been developed. Nomenclature initially suggested for these proteins was based upon their electrophoretic behavior under a given set of conditions. Because of increased interest in milk proteins of species other than bovine, the Committee suggests that these be identified as homologs of those already characterized in European, Bos taurus , and Indian, Bos indicus , cattle. Guidelines are given to aid in determining if homology exists. Provisional nomenclature is suggested for use in the interim until homology can be established.