[Recent developments in selected areas of food biotechnology]. / Neuere Entwicklungen auf ausgewählten Gebieten der Lebensmittelbiotechnologie.

With respect to literature and own experiments a review is given of the current international knowledge in special fields of food biotechnology. In this context nutritionally and physiologically relevant problems find particular consideration. After an introduction the development in technology, genetic engineering and protein technology is discussed. Then the following topics are dealt with: Single cell proteins (mushrooms, yeasts, bacteria, fungi, phototrophic microorganisms), soluble enzymes, enzyme inhibitors, immobilized systems (enzymes, cells, new processes and products), amino acids, sweeteners, microbial polysaccharides as hydrocolloids, organic edible acids, plant cell and tissue cultures and flavourings (enzymes, microorganisms, plant systems).

Influence of culture conditions on mycelial structure and polygalacturonase synthesis of Aspergillus niger.

Biosynthesis of polygalacturonase (PG) by A. niger strain R 1/214 correlates with the morphology of mycelium in submerged culture. The mean specific PG-synthesis (PG-U.g-1.h-1) increases with the degree of compactness of mycelium. PG-production can be optimized by a precise adjustment of the culture conditions after direct spore inoculation (diffuse mycelium) but the high synthesis as by compact mycelium is never obtained. Different reasons for the higher enzyme production by the pellet mycelium are discussed. PG-synthesis is assumed to be strictly connected with a limitation of nutrient and oxygen supply.

Aroma retention in modern bread production.

In highly industrialized countries bread consumption rate decreases in favour of flavour-intensive foods like meat, sweets, cocoa products or alcoholic beverages. This trend which is unfavourable from the nutritional point of view may be forced by flavour losses in modern bread technology itself. Sensory profile studies indicate 4 points which may change the traditional bread flavour, i.e. reduction of flour extraction rate, changes within bread formula, shortening of the dough procedures or of baking time. Possible ways for improving bread flavour in modern technology are discussed. The use of precursor systems as well as of biotechnical procedures deliver chances for future bread production with improved flavour quality.

[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.

[Physiological and cell biological characterization of the protease producer Thermoactinomyces vulgaris during prolonged culture in a stirred fermenter]. / Physiologische und zellbiologische Charakterisierung des Proteasebildners Thermoactinomyces vulgaris während Langzeit-Kultivation im Rührfermentor.

The physiological behaviour of Thermoactinomyces vulgaris - producing a thermostable serine-protease - was analyzed during fermentation. During 38 h the consumption of nutrients and oxygen as well as the rates of macromolecular and protease synthesis were measured. The morphological and ultrastructural changes of the mycelia were also studied. The mycelia grew exponentially for about 5 hours. After a short lag and a second slower growth phase, growth continued about linearly until the end, as was indicated by a constant rate of incorporation of labelled thymidine. However, at the same time a changing portion of hyphae - up to 45% - underwent lysis. According to the changing ratio of growing and lysing material, regarding the physiological activity of the culture the fermentation process could be divided into 4 periods. The formation of the protease started at the transition to the slow growth phase and continued linearly. The ability to produce the protease was attributed to a mycelium being formed after the shift down caused by limitation of supply of utilizable nitrogen compounds. The end of protease production 10 h later was correlated to a drastic decrease of the respiratory activity of the mycelia, probably caused by exhaustion of easily utilizable carbohydrates.

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.