[Analysis of gangliosides with special reference to milk]. / Analytik von Gangliosiden unter besonderer Berücksichtigung der Milch.

Gangliosides are sialic acid containing lipid compounds of a very complex nature, which are involved in many physiological processes. Gangliosides are very important constituents of membrane material in animal tissue, where they occur at very low concentrations. Improved analytical procedures are therefore required to study their behaviour, their role in living systems and their isolation for further investigations. In this paper, all analytical methods which may be important in the analysis of gangliosides are summarized and discussed. These include extraction, purification, enrichment and chromatographic procedures. In bovine milk, three gangliosides of known structure are present. Their isolation, structure and analytical importance (buttermilk) are also reported.

[A rapid computer-assisted method for the determination of urea in dairy products: skim milk powder from various sources]. / Eline rechnergesteuerte Schnellmethode zur Bestimmung von Harnstoff in Milcherzeugnissen: magermilchpulver unterschiedlicher Provenienz.

The enzymatic determination of urea in spray-dried skim milk powders was transferred to a computer aided photometric analysing system. Sample preparation is optimal using ultrafiltration, but the direct use of a milk sample is also possible with this system. The precision including sample preparation as coefficient of variation was about 0.69%, the standard deviation of the assay was 0.115-0.135 at an average content of 5.4-5.5 mg urea per 100 ml skim milk. The method was tested on 147 spray-dried skim milk powders, which were of varying origin and collected at different times of the year (Jan.-Oct.). In this way a representative survey for the urea content of a great number of skim milk powders was possible. It was also shown, that the urea content is highly affected by season. It is very constant from February to May (0.25-0.26%), and rises from June to October up to nearly 0.32%.

[Ion chromatographic determination of lactulose and epilactose in the presence of lactose and other carbohydrates in milk and milk products]. / Ionenchromatographische Bestimmung von Lactulose und Epilactose neben Lactose und anderen Kohlenhydraten in Milch und Milchprodukten.

A method is described for the analysis of carbohydrates in milk and milk products which develop during heat treatment. It is based on the separation of their borate complexes by anion exchange chromatography using two optimized buffer systems. A reliable qualitative and quantitative determination of the disaccharides lactulose and epilactose is possible even in the presence of large amounts of lactose (buffer system I). Using the second buffer system lactulose can also be detected in the presence of fructose. In addition other relevant carbohydrate components like ribose, mannose, galactose and glucose can be determined simultaneously.

New aspects of naturally occurring proteases in bovine milk.

The scientific literature on milk proteases, along with recent findings in the author's laboratory, are summarized and reviewed comprehensively. Emphasis is on detection of proteolytic enzymes and their activity, purification and kinetic characterization of the isolated enzymes, and technological problems associated with proteolytic enzymes in milk and milk products. Two serine proteinases isolated from milk are compared with plasmin of bovine blood serum. Results from these comparisons strongly suggest that milk proteinase I and plasmin are identical. Proteolysis studies with cold stored milk indicate a direct relationship between gamma-casein formation and milk proteinase association with casein micelles.

The formation of gamma-caseins during cooling of raw milk.

It has been shown that there is a time-dependent transfer of beta-casein and the milk serine proteinase system from micelles to milk serum with change of temperature from 38 to 4 degrees C. It has been established that the gamma-caseins can be formed by proteolytic degradation of beta-casein. By a simple extraction technique, the very hydrophobic gamma-casein fraction was separated from stored milks (26 and 4 degrees C) and estimated quantitatively. The results showed that the proteolytic degradation of beta-casein is faster at 4 degrees C than at room temperature and this can be explained by the immobilization of enzyme and substrate at the micelle surface at higher temperatures (26 degrees C). The results indicate that irreversible changes during cooling for short periods do not cause problems in milk processing, but the formation of gamma-caseins and phosphopeptides may influence the technological properties of raw milk stored for more than 48 h.

Model proteolysis of beta-casein by immobilized trypsin.

The enzymic degradation of beta-casein by immobilized trypsin is described and its use as a model system to study proteolytic reactions which occur in milk during processing is examined. Quantitative determination of proteolysis can be obtained by densitometric transmission measurement of patterns obtained by polyacrylamide-gel electrophoresis. Staining properties of some of the individual components in the electrophoretic patterns were determined by calibration with pure protein and peptide preparations. The isolation of pure beta- and gamma-caseins and the preparation of several forms of immobilized trypsin are outlined.

[A quantitative micromethod for the caffeine determination (author's transl)]. / Mikromethode zur quantitativen Bestimmung von Coffein.

A combined procedure with thin-layer-chromatography and densitometry is described for the quantitative estimation of caffeine in biological material. This method is applicable in the nanogram range. Test samples of less than 100 microliter may be used. The samples (capillary-blood) are extracted with the same volume of chloroform. Caffeine is separated from interfering compounds by thinlayer-chromatography. Commercial silica-60-plates with chloroform/acetone (9+1; v/v) as solvent are used. The running time is about 30 min. The quantitative densitometric determinations are performed in the remission mode at 273 nm. In the range from 10 to 60 ng/spot the calibration curve is linear. Accurate quantitative data will be obtained even at concentrations of 1 mg/l caffeine. The detection limit is at about 0.1 mg/l.

Gel diffusion--a simple and sensitive technique for the assay of proteinase inhibitors and its use for the determination of the ratio of proteinases in mixtures.

In casein-containing agarose gels, pepsin and chymosin form radial diffusion zones; the diameters of these zones show rectilinear correlations with the logarithm of the enzyme concentration at constant time. The sensitivity for both enzymes is below 1 microgram. Addition of the inhibitor pepstatin A to these enzymes causes a reduction of the diameters of the diffusion zones, with large differences for both the enzymes. With this procedure, the pepsin/chymosin ratio in rennet preparations was assayed with an accuracy of +/- 5%. Identification of the inhibitors allows the determination of amounts in the namomole range. This method is a simple technique for the evaluation of proteinases and their inhibitors in screening systems.

Heat induced crosslinks in milk proteins and consequences for the milk system.

The protein system of milk is rather unusual, there are nearly no interchain crosslinks found. Even intrachain crosslinks, especially disulfide bridges, are present only in about every fourth protein molecule. Heating causes dramatic changes in the structure of milk proteins, resulting in the formation of polymeric networks. The contribution of individual milk proteins, namely the beta-lactoglobulins, alpha-lactalbumin and chi-casein, to the formation of crosslinks is studied with respect to heating temperature and time, pH and atmosphere. Measured are changes in molecular weights and in the SH/SS-levels as well as the formation of dehydroalanine, lysinoalanine, lanthionine and isopeptide bonds. Some practical aspects of crosslinking in milk proteins are discussed.

[Nepsilon-(beta-Aspartyl)lysine and Nepsilon-(gamma-glutamyl)lysine crosslinkings in bovine colostrum (author's transl)]. / Vorkommen der Isopeptidbindungen Nepsilon-(beta-Aspartyl)lysin und Nepsilon-(gamma-glutamyl)lysin in Rindercolostrum

In bovine colostrum which was defatted and totally hydrolyzed by proteolytic enzymes, the isopeptides Asp Lys and Glu Lys were identified by ion-exchange chromatography and preparative separation. Isopeptides were identified in two of seven samples of colostrum. It is still to be clarified whether there is a correlation between the amound of gamma-glutamyltransferase and the appearance of these crosslinkings.

[Investigations on the production of mycotoxins and their quantitative evaluation. I. The production of penicillic acid by Penicillium cyclopium (author's transl)]. / Untersuchungen zur Bildung von Mykotoxinen und deren quantitative Bestimmung. I. Die Bildung von Penicillinsäure durch Penicillium cyclopium

A convenient thin-layer chromatographic screening procedure for the analysis of mycotoxins produced by Penicillium cyclopium is described. The production of penicillic acid is followed during a growth period of 44 days at two different temperatures. Quantitative evaluation is performed by UV densitometry at 234 nm. A biological profile is recorded showing the different ratios of four metabolites produced by Penicillium cyclopium and the definite effect of the growth temperature on the formation of penicillic acid in relation to the other metabolites.

[Polysaccharide additives in foods rich in proteins II. A simple, semi-quantitative technique of estimation (author's transl)]. / Polysaccharidzusätze in eiweissreichen Lebensmitteln. II. Eine einfache, semiquantitative Bestimmungstechnik

Samples of foods rich in protein, after removal of lipids and of proteins by enzymatic hydrolysis, are partially demineralized and concentrated. In the concentrate polysaccharides are detected and semi-quantitively estimated by performing the precipitations in graduated tubes, which are especially suitable for this purpose, and the precipitates taken to comparable densities by centrifugation under standard conditions.