Showing papers on "Lipase published in 1973"
TL;DR: A novel method for determining serum triglycerides, in which an enzymatic hydrolysis replaces the more commonly used saponification procedure, which is simple, rapid, and requires only 50 µl or less of sample.
Abstract: We describe a novel method for determining serum triglycerides, in which an enzymatic hydrolysis replaces the more commonly used saponification procedure. Under the conditions of the assay, the enzymatic hydrolysis can be completed in less than 10 min by the combined action of a microbial lipase and a protease. We have been able to demonstrate complete hydrolysis of triglycerides by thin-layer chromatography of the reaction products, by recovery of glycerol from sera of known triglycerides content, and by comparison of triglyceride assays on a number of sera assayed by our method vs. the AutoAnalyzer procedure. The hydrolysis is directly coupled to the enzymatic determination of glycerol, and is followed through absorbance changes at 340 nm. The assay is simple, rapid, and requires only 50 µl or less of sample. Because the enzymes used do not release glycerol from other compounds in serum, the hydrolysis can be considered specific for triglycerides.
3,113 citations
319 citations
TL;DR: The effect of conjugated bile salts on the activity of pancreatic lipase depends on their concentration in relation to their critical micellar concentration, and it appears justified to classify co-lipase as a co-enzyme for lipase.
Abstract: The effect of conjugated bile salts on the activity of pancreatic lipase depends on their concentration in relation to their critical micellar concentration.
Below the critical micellar concentration, conjugated bile salts slightly stimulate the initial rate of hydrolysis of tributyrine, an effect that may be caused by a protection of the enzyme from inactivation at the substrate-water interface.
Above the critical micellar concentration, conjugated bile salts almost completely inhibit lipase. The inhibition is more marked in alkaline reactions resulting in a pH optimum shift with increasing bile salt concentration. Bile salt inhibition of lipase is related both to the concentration of bile salt and to the substrate concentration or rather to substrate surface area, and most probably is complete when the interface is saturated with detergent.
Co-lipase, in the absence of bile salts, stimulates the activity of lipase, 1.3 to 1.4-fold in the whole pH range of its activity. Co-lipase overcomes the inhibition of lipase caused by bile salts with a shift in the pH optimum to 6–7 compared to 8–9 for lipase alone.
The different conjugated bile salts have similar effects, consideration being taken to differences in their critical micellar concentration; free bile salts have a less inhibitory effect on lipase and the stimulation by co-lipase shows no pH optimum shift.
Detergents of the acyltaurine type such as decanoyltaurine and dodecanoylsarcosyltaurine inhibit lipase in a similar manner to the conjugated bile salts and this inhibition is also released by co-lipase. Detergents such as dodecylsulphate above the micellar concentration, irreversibly inhibit lipase. The simultaneous presence of bile salts protects the enzyme from being irreversibly inactivated.
Lipase and co-lipase interact in a stoichiometrical relationship and it appears justified to classify co-lipase as a co-enzyme for lipase.
237 citations
TL;DR: The serous glands of rat tongue were found to contain a potent lipolytic enzyme which hydrolyzed triglyceride to mostly diglyceride and free fatty acids (FFA) at pH 4.5-5.4 and it is proposed that this reaction is the first step in the digestion of dietary lipid.
Abstract: The serous glands of rat tongue were found to contain a potent lipolytic enzyme which hydrolyzed triglyceride to mostly diglyceride and free fatty acids (FFA) at pH 4.5-5.4. Homogenates of lingual serous glands from adult rats hydrolyzed 40-70 mmol of triglyceride/g per h. The soft palate, anterior oral pharyngeal wall, and lateral oral pharyngeal glands also contained the activity, but at a much lower level. The lipolytic activity was also found in saliva collected through an esophageal cannula and in stomach contents of rats fed a fat-rich meal. The stomach contained very little activity, however, when saliva was excluded. Lipolytic activity was not found in the stomach wall or in the parotid, submandibular, and sublingual glands. The findings suggest that the lingual serous glands secrete a lipase which catalyzes in the stomach the conversion of triglyceride to partial glycerides and FFA. It is proposed that this reaction is the first step in the digestion of dietary lipid.
232 citations
145 citations
TL;DR: Post-heparin plasma in the rat contains triglyceride lipase activity of both hepatic and extrahepatic origin, and a 360-fold enhancement of plasma membranelipase activity over that in whole liver homogenate was achieved by heparin affinity chromatography.
128 citations
TL;DR: Analysis of products enzymatically derived from labeled cutin by thin-layer chromatography and radio gas-liquid chromatography showed that the Fusarium enzyme released all classes of cutin monomers.
105 citations
TL;DR: Two co-lipases have been isolated from extracts of porcine pancreatic gland and contain the same number of acidic and basic amino acids and therefore most probably only differ in the extent of amidation of their glutamic and aspartic acid residues.
97 citations
TL;DR: In contrast with the statement of these authors that an accurate calibration of the method—conversion of ΔA into μmoles of fatty acid liberated per minute—is not possible, a relatively simple way for such a conversion is discovered.
89 citations
TL;DR: Two colipases preventing lipase inhibition by bile salts have been purified with a 30% yield from defatted porcine pancreas and ultracentrifugation assays were consistent with a bile salt-mediated conversion of colipase into another form possessing a higher MW and the ability to give with lipase a biles salt-resistant complex.
89 citations
TL;DR: It is concluded that the enzyme has a rather low substrate specificity and that the presence of activating serum factors is not necessary for catalysis to occur.
TL;DR: It would appear that levels of pancreatic lipase are increased when the fat content of the diet is raised from about 5% to 15-22%, but that little or no additional increase in lipase levels can be attained by any further increase in the amount of dietary fat.
TL;DR: In this article, the authors used ammonium sulfate fractionation, DEAE-Sephadex column chromatography, gel-filtration, and column-clustering to obtain a homogeneous solution of Geotrichum candidum link.
Abstract: Lipase (EC 3. 1. 1. 3) of Geotrichum candidum Link was purified by means of ammonium sulfate fractionation, DEAE-Sephadex column chromatography, gel-filtration on Sephadex G-100 and Sephadex G-200, and was finally crystallized in concentrated aqueous solution. It was confirmed that the crystallized preparation was homogeneous electrophoretically and ultracentrifugally. It was estimated with the crystalline enzyme that the sedimentation constant (s20, W) was 4.0, the isoelectric point was pH 4.33, and the molecular weight was 53, 000_??_55, 000. From the result of amino acid analysis, none of sulfur containing amino acid was detected in the enzyme. It was also recognized that the crystalline preparation contained about 7% of the carbohydrate and very small amount of lipid. It was characterized that the lipase was the most active at pH 5.6_??_7.0 on olive oil, at 40°C and was stable in the range of pH 4.2 to 9.8 at 30°C for 24 hr, and was stable below 55°C for 15min.
Patent•
30 May 1973TL;DR: In this paper, a glycerol detection system was proposed by using enzymes in the presence of carboxylesterase and an alkali metal or alkaline earth metal alkyl sulfate.
Abstract: Triglycerides are determined by enzymatic saponification with lipase and measurement of the liberated glycerol by carrying out the saponification in the presence of carboxylesterase and an alkali metal or alkaline earth metal alkyl sulfate; reagents are provided for carrying out the process and comprise a buffer and, as a saponification agent, lipase, carboxylesterase and the alkali metal or alkaline earth metal alkyl sulfate, and a glycerol detection system.
TL;DR: A simple two‐parameter model is developed describing the degree of repression or induction in fermentation media.
Abstract: The synthesis of extracellular enzymes by microorganisms frequently occurs under genetic control. A simple two-parameter model is developed describing the degree of repression or induction in fermentation media.
The case of substrate utilization by an extracellular enzyme was analyzed for a vegetable oil-lipase-yeast system. It is shown that fatty acids released by the lipase may accumulate in the early stage of growth and exert an influence on the limiting after which relatively little repression or induction takes place.
Expressions are also derived for growth and extracellular enzyme synthesis in single-and multistage continuous cultures. When the cells grow on a directly available soluble substrate, the specific enzyme synthesis is maximal at low dilution rates in the case of repression and at high dilution rates in the case of induction. If the substrate is not directly available, a single continuous stirred tank reactor stage may not be sufficient for efficient substrate utilization; for fermentation processes where an insoluble has to be broken down before the cells can assimilate it, a plug flow type fermentor rather than a mixed chemostat may prove more satisfactory.
TL;DR: The incorporation of delipidated very low density lipoprotein protein with the lipid substrate increased the maximal reaction velocity of lipase activity at all hydrogen ion concentrations, and also decreased the apparent Michaelis constant of the reaction at alkaline pH.
TL;DR: Combined guinea-pig cortex and cerebellum was shown to contain triglyceride lipase, diglyceridelipase and monoglyceride Lipase, which were assayed by the release of [1-(14)C]palmitate from [1-C]Palmitoylglycerol esters, which indicated that aging of membranes at 0 degrees C decreased activity, which could still be stimulated by noradrenaline.
Abstract: 1. Combined guinea-pig cortex and cerebellum was shown to contain triglyceride lipase, diglyceride lipase and monoglyceride lipase, which were assayed by the release of [1-(14)C]palmitate from [1-(14)C]palmitoylglycerol esters. Triglyceride lipase and diglyceride lipase were found in all particulate fractions. 2. With osmotically ruptured synaptosomes the rates of release of palmitate from glyceryl tripalmitate and glyceryl dipalmitate were 7-25mumol/h per g of protein and 0.18-0.69mmol/h per g of protein respectively. The logarithm of the rate of hydrolysis of glyceryl monopalmitate increased linearly with the logarithm of protein concentration. The pH optima of triglyceride lipase and diglyceride lipase were between 7 and 8. The pH optimum for monoglyceride lipase was approx. 8. 3. Triglyceride lipase and diglyceride lipase of osmotically ruptured synaptosomes were stimulated by noradrenaline, 5-hydroxytryptamine and adrenaline. Triglyceride lipase of isolated synaptic membranes was stimulated by 0.01-1mm-noradrenaline. Aging of membranes at 0 degrees C decreased activity, which could still be stimulated by noradrenaline. Diglyceride lipase of isolated membranes was stimulated by 1mum-1mm-noradrenaline. The activity of triglyceride lipase in isolated synaptic vesicles was diminished by 1mm-5-hydroxytryptamine.
TL;DR: A nephelometric method for the determination of serum lipase is described, which correlates with and is a more sensitive indicator of pancreatitis than the Cherry-Crandall method.
TL;DR: It is suggested that similar hydrophobic heads and hydrophilic tails and asymmetric charge distributions establish the orientation of many enzymes which act at interfaces, and many phospholipases, for instance, appear to be charge-oriented, and the carbohydrate residues of ribonucleases and many other glycoproteins may be hydphilic tails.
Patent•
25 Jul 1973TL;DR: In this article, the lipase-plus-activator compositions are utilized to pre-soak soiled fabrics, which are subsequently laundered using conventional household equipment, such as washing machines.
Abstract: Compositions and methods for removing oily triglyceride stains from fabrics employing a lipase enzyme and a lipase activator selected from the group consisting of naphthalene sulfonates, certain polyoxyalkylene derivatives of ethylenediamine and certain acylamino acid salts. The lipase-plus-activator compositions are utilized to pre-soak soiled fabrics, which are subsequently laundered using conventional household equipment.
TL;DR: An accurate and convenient colorimetric method for the determination of serum and urinary lipase is described, which is rapid, sensitive, uses small quantities of serum, and is particularly suitable for batch analysis.
TL;DR: Data presented indicate that adding sodium glycocholate prevents inactivation of lipase and does not lead to activation of this enzyme, and Comparative studies show that Triolein is the superior substrate for lipase, but olive oil can be substituted for triolein for routine clinical laboratory use.
Abstract: A method is presented for measuring lipase activity in serum by use of either triolein or olive oil as substrate. The method requires 3 to 8 min for each measurement and 5 to 8 min for initial set-up. The reaction mixture is maintained at the optimum pH of 8.8 by automatic addition of sodium hydroxide (10 mmol/liter) with a " pH-Stat." Amount of sodium hydroxide delivered per unit time reflects lipase activity. When reaction conditions—such as pH, substrate concentration, emulsifier concentration, and type and concentration of added bile salts—were made optimal, the reaction kinetics were zero order. Data presented indicate that adding sodium glycocholate prevents inactivation of lipase and does not lead to activation of this enzyme. Comparative studies, in which triolein and olive oil, respectively, are used as substrates show that triolein is the superior substrate for lipase, but olive oil can be substituted for triolein for routine clinical laboratory use. The suggested upper limit of normal is 200 and 160 U/liter for triolein and olive oil substrates, respectively.
TL;DR: The pH dependency of the rate constant for dioctanoin hydrolysis reveals an ionizable basic group at the active site of the enzyme with pKa of 6.38, consistent with fully hydrated enzyme molecules acting upon substrate molecules lying within the insoluble mono-layer.
TL;DR: The enzyme showed the optimum pH between 7.5 and 8.0, and the optimum temperature at about 37°C, but completely lost the activity by heating 60°C for 15 min this paper.
Abstract: The enzyme showed the optimum pH between 7.5 and 8.0, and the optimum temperature at about 37°C. It was stable over the pH range from 4 to 9 and below 40°C, but completely lost the activity by heating 60°C for 15 min. The enzyme was activated by low concentration of calcium ion but inhibited partially by high concentration of calcium ion and by EDTA. With respect to substrate specificity, the enzyme exhibited a high specificity toward triglycerides and hydrolyzed ester bonds of short-carbon chain triglycerides faster than long-carbon chain triglycerides, whereas it catalyzed the hydrolysis of the oils from rice bran, olive and coconut. When 2-oleo-1,3-distearin was used as substrate, the enzyme was capable of preferentially hydrolyzing fatty acid ester bonds at the 1,3-position.
TL;DR: It was recognized that Geotrichum candidum Link which was selected as the efficient lipase producer formed lipase only in the presence of substrate or its relating compounds such as oils or fatty acids in a cultivation medium.
Abstract: It was recognized that Geotrichum candidum Link which was selected as the efficient lipase producer formed lipase only in the presence of substrate or its relating compounds such as oils or fatty acids in a cultivation medium. From the experimental results obtained by the cultivation of the microorganism and also by using of washed cells, it seemed that lipase was formed inducibly. It is likely that the produced lipase is localized around the cell wall and membrane and it is released from the cells after a certain period from the inducible synthesis.
TL;DR: A Langmuir trough was converted into a fully automatic recording bidimensional barostat which was used for investigating in detail the effect of suface pressure on the digestion of various ester monolayers by pure pancreatic lipase (glycerol-ester hydrolase).
TL;DR: The results indicate that L cells in tissue culture can utilize serum triglycerides and suggest that the predominant mechanism of uptake involves the intact molecule and does not require prior hydrolysis.
TL;DR: American lobsters, Homarus americanus, were fed diets containing 0, 5, and 20% starch for periods up to 37 days, and specific activities and absolute amounts of proteinase, lipase and, to a lesser extent, amylase were found to rise on prolonged feeding.
Abstract: American lobsters, Homarus americanus, were fed diets containing 0, 5, and 20% starch for periods up to 37 days. Digestive enzymes in the stomach juices of these and fasted lobsters were assayed. The base-line, or true fasting enzyme level was considered to be those values found after feeding for 2 days on a 5% starch diet. Compared to this basal level both specific activities and absolute amounts of proteinase, lipase and, to a lesser extent, amylase were found to rise on prolonged feeding. Further feeding (all diets) allowed products of a greater proportion of proteinase than amylase or lipase; alteration of the starch content of the diet had no consistent effect.
TL;DR: Olive oil hydrolyzing fraction (F-3) of Mucor lipase was separated into two fractions by means of CM-Sephadex column chromatography as mentioned in this paper.
Abstract: Olive oil hydrolyzing fraction (F-3) of Mucor lipase was separated into two fractions, F-3A and F-3B, by means of CM-Sephadex column chromatography. Both fractions were homogeneous and F-3A was crystallized. The pH optimum for olive oil hydrolysis of F-3A was at 9.0 and that of F-3B was at 8.0.Chain specificities of the both enzymes were different with each other. Sedimentation coefficient (S20, w) of F-3A was 2.8 S and that of F-3B was 3.1 S.Molecular weights calculated from sedimentation equilibrium data were 25,400 for F-3A and 29,000 for F-3B.
TL;DR: It is concluded that lipoprotein lipase activities from heart and liver are catalysed by different enzymes and that the heart enzyme contributes not more than 5% to the overall lipase activity in postheparin serum.