Purification, characterization and kinetic properties of the rabbit gastric lipase.

Rabbit gastric lipase was purified from an acetonic powder of rabbit stomach fundus. 25 mg of pure rabbit gastric lipase (glycerol ester hydrolase, EC 3.1.1.3) was obtained from 30 rabbit stomachs after ammonium sulfate fractionation, Sephadex G-100 gel filtration and cation exchange (mono S column) using a fast protein liquid chromatography (FPLC) system. The pure enzyme obtained was resistant to acidic pH conditions, and had specific activities of 1200, 850 and 280 U/mg, using, respectively, short- (tributyroylglycerol (TC4)), medium- (trioctanoyl- to tridecanoylglycerol (TC8-TC10)) and long-chain (soybean oil) triacylglycerols. The amino-acid composition was determined, and the first 30 N-terminal amino-acid residues were sequenced. Interfacial denaturation and catalytic properties on triacylglycerol emulsions were studied. Rabbit gastric lipase turned out to be structurally and kinetically very similar to human gastric lipase.

Screening of preduodenal lipases in several mammals.

The tissular localization of preduodenal lipases was studied from the tongue to the pyloric portion of the stomach in 11 mammals. Lipolytic activities were clearly differentiated from those of pancreas. All lipase activities show an acidic pH optimum, except the gastric enzyme from hog. For every mammal tested, preduodenal lipase activity was associated mainly with only a single tissue located either in tongue, or in the pharyngeal area, or in the stomach. Resistance to acidic pH medium allows the classification of lipase activities into three groups. These results are related to the dietary habits and zoologic classification of the different animal species.

Titrimetric assay of pancreatic lipase in pharmaceutical preparations.

Lyophilized pancreas and pancreatic extracts are widely used in therapy of pancreatic exocrine function deficiencies. Among the measures for quality of the extracts, assay for lipolytic activity appears to be one of the best. However, assay precision is poor, because the specificity and mode of action of lipase requires careful optimization of the assay parameters, especially substrate and measurement conditions. In the method proposed here, substrate quality is improved by the use of sodium desoxycholate and a high-speed stirrer for more reproducible emulsions. Fatty acids are assayed by a titrimetric method, using an electronically monitored pH meter. A comparative statistical study of the U.S. Pharmacopeia method, the International Pharmaceutical Federation (FIP) method, and a modified FIP method showed the latter to have improved accuracy and reproducibility.

The structure of sialyl-glycopeptides of the O-glycosidic type, isolated from sialidosis (mucolipidosis I) urine.

Sialyl-glycopeptides containing an O-glycosidically linked tetrasaccharide chain were obtained from the urine of a patient suffering from mucolipidosis I. Isolation of these compounds was achieved by gel filtration, ion-exchange chromatography and preparative paper chromatography. Their structures were determined by a combination of carbohydrate and amino acid analysis, dansylation, periodate oxidation, methylation studies, enzymatic hydrolysis and 1H-NMR spectroscopy, to be as follows: (formula; see text) wherein R = peptide linked through -Thr-, -Ser-Thr- or -Thr-Ser-. The finding of these glycopeptides in urine shows that mucolipidosis I is characterized by a general "glycoprotein-specific" sialidase deficiency. The possibility of the existence of a human endo-alpha-N-acetylgalactosaminidase is discussed.

[Excretion of sialoglycopeptides in mucolipidosis I]. / Excrétion de sialoglycopeptides, associée à la mucolipidose de type I.

Mucolipidosis I involves a tremendous increase of the urinary excretion of sialoglycopeptides and sialyloligosaccharides. This enhancement is due to the excretion of O- and N-glycosidic peptides and oligosaccharides, which normal urine is devoid of as shown by the chemical composition and electrophoresis or thin-layer analysis. This finding is in agreement with the recent finding of an alpha-neuraminidase deficiency for this disease.