[Nutritional physiology of whey and whey components]. / Ernährungsphysiologische Bedeutung von Molke und Molkenbestandteilen.

Deriving from positive effects of whey drinking cures in antiquity, the Middle Ages and modern time, a review is given on nutritional significance of whey. The proteins are essential components of whey and belong to the proteins with highest biological value because of their amino acid composition. Besides, they show fundamental functional properties, which enable a varied application in foods, dietetic foods and beverages in form of different whey products (powder, protein concentrates and isolates). Whey proteins have found considerable usage in infant's nutrition as whey predominant formulas as well as whey protein hydrolysates in case of cow's milk protein intolerances. A recent field of research are biological active peptide sequences which become effective during digestion and are of importance for secretion of entero hormones as well as for immune enhancing effects. They may contribute to assess the biological value of whey proteins under enlarged points of view and to develop new application forms and areas. It is pointed to further fields of application (e.g. adipositas, gout, kidney insufficiency). Concerning the quantitatively most dominant lactose in whey, it is dealt with its importance for the healthy development of infants (adaptation to the increased lactose content of mother's milk) as well as with lactose intolerance and galactosaemia. In case of mineral salts of whey it is emphasized the high nutrient density of calcium (prophylaxis for osteoporosis), the beneficial Ca:P and Na:K proportions (antihypertensive in case of the last one), the promotion of absorption of mineral salts by lactose, and the high content of iodine. The whey is rich in B-vitamins, which contribute essentially for their satisfaction or requirement in case of a corresponding consumption. To be emphasized is the vitamin B12 in milk and whey, which is the sole source of this indispensable nutrient for blood-formation and cell division in lacto-ovo-vegetarian nutrition. In conclusion, a summarizing dietetics valuation of whey is performed.

[Importance of peptides from enzymatically degraded proteins as components of foodstuffs]. / Bedeutung von Peptiden enzymatisch abgebauter Proteine als Bestandteile von Nahrungsmitteln.

In comparison with proteins, peptides occur only in very small amounts in foods. An exception are peptides, which result from proteins to a higher or lower extent in the course of natural or microbial disintegration or ripening processes. Recently, proteinases are used for process optimization or quality improvement in the production or processing of foods. Usually, the protein degradation in these cases is only small, but functional properties may be improved. Peptides frequently show a bitter taste and may limit the sensory value of foods. Sources and possibilities to remove the bitter taste are discussed. In dietetics protein-free peptide diets are introduced for special indications. They contain oligopeptides which are produced by intensive hydrolysis of proteins. The intestinal absorption of small peptides is different from that of free amino acids. Finally, physiological effects of some dietary peptides are discussed (e.g. peptides as enzyme inhibitors, coeliac disease as a gliadine-peptide induced enteropathy, exorphines as neurotransmitters).

[Intracellular proteases of Thermoactinomyces vulgaris. 1. Occurrence of the proteases in the cell and dynamics of their formation during cultivation]. / Intrazelluläre Proteasen aus Thermoactinomyces vulgaris. 1. Mitt. Vorkommen der Proteasen in der Zelle und Dynamik ihrer Bildung im Verlauf der Kulturführung.

The investigation concerning the localization of proteolytically active enzymes in the cells of Thermoactinomyces vulgaris show that the enzymes are present in a dissolved form in the cellular extract (75%) as well as linked to the solid cell components (25%). The ratio of the activities of the soluble periplasmatical fraction to the soluble cytoplasmatical fraction to the insoluble cytoplasmatical membrane fraction was found to be 2:1:1. The formation of the proteins during the cultivation is measured by detecting the activity in the cellular extract. As soon as enough of biomass is present (weighable) in the medium, the protease activity can be detected in the cellular extract. It increases during the fermentation and amounts to 1 to 2% of the activity of the proteases in the medium between the 10th and the 24th hour. A correlation between the formation of biomass and enzymes does not exist.

[Intracellular proteases of Thermoactinomyces vulgaris. 2. Biochemical characterization of proteases in cell extracts]. / Intrazelluläre Proteasen aus Thermoactinomyces vulgaris. 2. Mitt. Biochemische Charakterisierung der Proteasen des Zellextraktes.

The intracellular proteases (IP) of the cellular extract of Thermoactinomyces vulgaris are characterized as to the biochemical properties compared with the corresponding extracellular proteases (EP). According to that, the storage stability and the temperature and pH behaviour (optimum, stability) of both of the proteases are identical. Nevertheless, differences were detected between IP and EP after the action of several effectors and different substrates. As could be seen after a column chromatographic separation of the IP of the cellular extract, it is composed of at least 3 proteases, two of them are serine proteases which can be inhibited moreover unspecifically by p-chloromercuri benzoate. The purified proteases (IP) are very instable and therefore not yet characterized in detail.

Preparation and characterization of proteases from Thermoactinomyces vulgaris. V. Investigations on autolysis and thermostability of the purified protease.

Thermitase, the main component of the proteases of the culture medium from Thermoactinomyces vulgaris, is degraded by autolyses (increase of liberated amino groups) and thereby inactivated especially at elevated temperature, at alkaline pH-values and in the absence of added substrates. As shown by polyacrylamide gel electrophoresis autolysis is an essential part during heat inactivation (complete disappearance of the thermitase band after heating the enzyme at 85 degrees C for 5 min). The quantitative comparison of autolysis and heat inactivation as well as the kinetics of reversible inhibition of the enzyme by HgCl2 at different temperatures showed that above 60 degrees C thermal denaturation of the enzyme protein contributes to thermitase inactivation. Ca2+-ions (20 mM) have a stabilizing effect against both autolysis and thermal denaturation (inactivation) of thermitase.

[Substrate specificity of a protease from Thermoactinomyces vulgaris]. / Untersuchungen zur Substratspezifität einer Protease aus Thermoactinomyces vulgaris.

The cleavage specificity of a protease from Thermoactinomyces vulgaris (thermitase) was determined by the insulin B-chain and the cleavability of casein and haemoglobin by this enzyme as compared to other proteases (trypsin, chymotrypsin, proteases from Bac. megaterium and cytophages). The most intense splitting effect on the substrates under investigation (insulin B-chain, casein and haemoglobin) is exerted by thermitase, i. e., the unspecificity of this enzyme is especially marked.

[Characterization of a protease from Thermoactinomyces vulgaris (thermitase). 1. Purification of thermitase]. / Charakterisierung einer Protease aus Thermoactinomyces vulgaris (Thermitase). 1. Untersuchungen zur Reinigung der Thermitase.

The paper deals with the purification of the microbial protease preparation "thermitase" (submerged cultivation of Thermoactinomyces vulgaris; treatment of culture filtrate with ethanol or Na2SO4, vacuum drying of precipitate). The crude substance was purified by column chromatography on Sephadex G-75, DEAE-Cellulose and Sephadex G-50. The proteolytically active fractions were in each case united, freeze dried and tested for protein components and protease activity by gel electrophoresis. After passage of the third column the isolated protease (4.5 fold enrichment in the specific activity) was further characterized. The electropherogram (pH 8.9) presented a protease band moving to the anode which was accompanied by 2 very weak protease bands. Furthermore there could be detected a very active protease band (main component of Thermitase) as well as a side band with lower activity both moving to the cathode. The freeze dried preparation contained 85% protein and 4% carbohydrates (glucose as single monomer component after acid hydrolysis). A molecular weight of 11,000 was determined by chromatography on Sephadex G-75. This value is critically discussed. Hints are given for autolytic processes taking place during the purification procedure.

[Characterization of a protease from Thermoactinomyces vulgaris (thermitase). 2. Single-step fine purification and protein-chemical characterization]. / Charakterisierung einer Protease aus Thermoactinomyces vulgaris (Thermitase). 2. Einschritt-Feinreinigung und proteinchemische Charakterisierung.

The fine purification of an alkaline protease (thermitase) from Thermoactinomyces vulgaris by means of isoelectrical focussing in the flat-bed procedure using granulated gel is reported. An Na2SO4-precipitated crude product serves as the starting material. Isoelectrical focussing leads in a single step to a highly purified protein with an uniform N-terminal end group. The enzyme has an IP at 9.0 and a mol. wt. of 37,400; it consists of a polypeptide chain with arginine as the N-terminal, and tyrosine as the C-terminal end group. In addition to an essential serine residue, a SH group could be demonstrated which is hardly accessible in the native enzyme. Furthermore, the influence of different protease inhibitors was studied.

[Characterization of a protease from Thermoactinomyces vulgaris (thermitase). 3. Substrate specificity and properties of partially purified thermitase]. / Charakterisierung einer Protease aus Thermoactinomyces vulgaris (Thermitase). 3. Substratspezifität und einige Eigenschaften der teilgereinigten Thermitase.

During the process of cultivation of Th. vulgaris several proteases are formed. In the present investigation the extensively purified major component was used. The substrate specificity was determined by means of 7 proteins, 7 amino acid esters, 5 fatty acid esters and 15 amino acid 4-nitroanilides. Among the protein substrates tested, urea denaturated hemoglobin was split best, followed by gelatin, casein, field bean protein, serum albumin and gluten. The weakest rate of hydrolysis was observed with elastin. In contrast to this acetyl-(L-ala)3-methylester, that is a substrate for elastase, was split best from all the esters tested. Only 8% of this activity could be found with the chymotrypsin substrates acetyl-L-tyr-ethylester and acetyl-L-phe-ethylester and 1% of the above activity with the trypsin substrates tosyl-L-arg-methylester and benzoyl-L-arg-methylester. The fatty acid esters and the p-nitroanilides were hydrolyzed much more slowly. The pH-optimum of thermitase was found in the weakly alkaline region of pH 7 to 9. There were only small differences between the individual high and low molecular substrates. The temperature optimum was between 60 and 75 degrees C for esters and p-nitroanilides as substrates and at 90 degrees C for casein. It should be mentioned that the enzyme was quickly inactivated at temperatures above 70 degrees C.

[Formation of bitter peptides in cheese and from casein]. / Zur Entstehung bitterer Peptide in Käse und aus Casein

Whereas a slightly bitter taste is desirable in certain foods, it is an off-flavour in cheese which may even lead to unfitness for consumption. Bitter principles from cheese have been found to be peptides with molecular weights ranging from 2000 to 3000. For the purpose of further characterization, bitter peptides were isolated from enzymatic casein hydrolysates as well as from bitter cheese and purified. 30 proteases from different origins proved to be able to form peptides with bitter taste of varying intensity from casein. Present experience shows that the formation of bitter peptides during casein hydrolysis can be inhibited only to a very small measure. Bitter peptides are extrmely resistant to proteases, which is probably attributable to their high contents of hydrophobic amino acids and hydrophobic bonds. The detection of only N- or C-terminal amino acid in each of 11 different bitter peptides shows that peptide chains are present and not cyclic peptides as repeatedly assumed. It must be aimed at avoiding the cheese defect "bitter" by using appropriate starter cultures and rennet substitutes as little disposed as possible to produce bitter peptides.