Ines Mandl

[92] Pancreatic elastase

Publisher Summary As the mechanism of action of elastase is not definitely known, all assay methods are based on measuring the amount of insoluble substrate solubilized in a certain time. The assay described can be used with crude preparations, as other pancreatic enzymes do not attack elastin. This procedure is preferred over gravimetric methods or others determining total protein solubilized, as elastin is often contaminated with small amounts of collagen which has been denatured and thus become susceptible to the action of trypsin and other proteolytic enzymes, including crude elastase. Whereas collagen is denatured very easily, elastin withstands heat and acids and alkalis of considerable strength. Any enzyme digesting elastin substrates is therefore an elastase, and the confusion that occurs with collagen substrates is not likely. On the other hand, crude extracts as well as crystalline and purified elastases show activity against unspecific protein substrates. New standard curves must be run with fresh batches of substrate. An inactive elastase precursor zymogen has been found in pancreas. It is activated by trypsin or enterokinase and is purified by adsorption on elastin.

Solubilization of insoluble matter in nature: I. The part played by salts of adenosinetriphosphate☆

Abstract The ability of soluble salts of ATP to solubilize numerous inorganic and organic compounds in neutral or slightly alkaline medium and to keep these compounds in solution is demonstrated. The latter can be natural products or synthetic substances. Difficulty soluble salts of ATP themselves are solubilized at physiological conditions of pH and temperature by other solubilizing agents. Among the latter are normal constituents of cells and products of intermediary metabolism. To these phenomena connected in different ways with ATP might be attributed essentiality as ATP is an omnicellular substance and plays a part in the course of many important biological processes.

Solubilization of insoluble matter in nature II. Part played by salts of organic and inorganic acids occuring in nature

Abstract It could shown that the salts of inorganic acids, and especially of a great number of orgaanic acids are capable of solubilizing insoluble mineral constituents and organic materials, or of preventing their precipitation. These compounds are formed from elements are formed from elements which belong to all groups of the periodic system and the most different classes. The solvents are found everywhere; they are obligatory intermediaries, continually reformed, or final products of metabolism, or cellular constituents. They perform at th same time the function of carrier of the solubulized material. The transformation products of materials of high molecular weight which can form salts are often excellent solvents. The general importance of these phenomena is discussed.

Characterization of some sugars of interest to biochemistry.

Summary Up till now readily formed derivatives from which the free sugars could be easily recovered were lacking in the case of several carbohydrates of biochemical interest. 2,5-Dichlorophenylhydrazones satisfy these requirements. Their preparation and characteristics are described. Asymmetrical diphenylhydrazine is preferable as a reagent for the separation of arabinose and ribose.

[22] Ketohexose phosphates

Publisher Summary Fructose-l,6-diphosphate (FDP) is most easily prepared from commercial bakers yeast. This eliminates the necessity of washing and pressing brewers yeast and yields satisfactory results with most brands. Fresh Atlantic, Federal, Blue Ribbon, and National Grain have been used successfully, but fresh Fleischmanns yeast cannot be used. Excellent results, however, were obtained with dried Fleischmanns as well as dried National Grain and Federal yeasts. Procedures for the use of both fresh and dried preparations are given by Neuberg and Lustig. Besides the usual determinations of organic P, colorimetric, spectrophotometric, and paper chromatographic methods of analysis is used. Fructose-l-phosphate (F-l-P) can be prepared by two different methods, both enzymatic, with FDP as the starting material. 1. Hydrolysis of FDP by bone phosphatase 21 gives a mixture of F-l-P, F-6-P, and aldose monophosphates. The latter can be removed by oxidation of the Ba salts with Br; the remaining fructose monophosphates are converted into their brucine salts and separated by fractional crystallization from water. 2. Enzymatic condensation of dihydroxyacetone-l-phosphate and D-glyceraldehyde by muscle or yeast aldolase, which should theoretically give a mixture of F-1-P and sorbose-l-phosphate, actually yields F-1-P only, almost quantitatively and irreversibly. In the presence of the enzyme, FDP, used as starting material, is converted to dihydroxyacetone phosphate and glyceraldehyde phosphate; the latter is in turn converted to dihydroxyacetone phosphate.

5-Phospho-d-arabonic acid

Abstract A simple method for the preparation of 5-phospho- d -arabonic acid by oxidation of d -fructose 6-phosphate is described. The significance of such an easily accessible phosphorylated product of the carbohydrate series obtainable in excellent yield is discussed.

[35] Preparation of d(+)-2-phosphoglyceric acid

Publisher Summary This chapter presents a procedure for the preparation of 2-phosphoglyceric acid (2PGA). 2-PGA can be prepared by two distinct methods; in both of them 2-PGA is isolated from the mother liquor remaining after precipitation of the less-soluble acid Ba salt of 3-PGA. The compound obtained from yeast maceration juice, when sugar fermented in the presence of fluoride. Neuberg accomplished total synthesis from D-glyceric acid and metaphosphoric acid ethyl ester. Isolation of pure D-2-PGA involves alcohol precipitation, which is repeated several times until the ratio of the optical rotation of the mixture no longer increases to the fermentation value. The final precipitate is then redissolved in the calculated amount of dilute HNO 3 and any phosphopyruvic acid is destroyed by adding a few milliliters of 5 % HgCl 2 and letting the solution stand for 20 minutes at room temperature. Hg is then precipitated with H 2 S, inorganic P with Mg mixture, and the precipitates filtered off. Procedure according to Neuberg include reaction of 26.5 g. of glyceric acid and 41 g. of metaphosphoric acid, which yields D(-)-3-PGA as the chief product together with a smaller amount of D(+)-2-PGA in a ratio of about 10:1. The isolation of 2-PGA from the remaining filtrate is based on the finding 4 that the Sr salt of 3-PGA is much less soluble in hot than in cold water.

[34] Preparation of d(−)-3-phosphoglyceric acid

Publisher Summary This chapter discusses the preparation of 3-phosphoglyceric acid (3-PGA). 3-phosphoglyceric acid (3-PGA) may be prepared by three different methods. It may be prepared biologically from yeast and sugar phosphate solution in the presence of acetaldehyde or another hydrogen acceptor and fluoride. The method discussed in this chapter has advantages of using commercial Bakers yeast instead of the more difficultly available washed and pressed bottom yeast. Racemic 2-PGA prepared from Na β-glycerophosphate can be converted by muscle extract to D(-)-3-PGA, which can be isolated in pure form, owing to the low solubility of the Ba salt of PGA compared to that of 2-PGA. Total synthesis has been accomplished from free D-glyceric acid and metaphosphoric acid ester by an adaptation of the method originally devised for the racemic compound. D(-)-3-PGA is the main product and is separated as the acid Ba salt from the D(+)-2-PGA formed at the same time.

Conversion of a phosphorylated ketohexose (d-fructose 6-phosphate) to a phosphorylated aldopentose derivative (d-arabonic acid 5-phosphate)☆

Abstract It is established that the product of oxidative degradation of d -fructose 6-phosphate is d -arabonic acid 5-phosphate.