Showing papers on "Amylase published in 1970"

Effects of Diet on Salivary Secretion and Composition

Abstract: The composition of saliva is potentially of great importance to the prevention of caries. Saliva contains buffers that tend to reduce the fall in pH that is associated with acid formation from carbohydrate by the dental plaque; it also contains urea, a substrate for base formation by dental plaque. Saliva contains at least one enzyme (amylase), which is involved in the first stages of the metabolism of starch, a component of many foods. In addition, saliva contains ions such as calcium, phosphate, and fluoride, the concentrations of which determine the pH at which tooth mineral will begin to dissolve. The main criticism that can be leveled against the majority of studies on the effects of diet on saliva is that the experimental design did not control the nondietary variables that can affect salivary flow rate or composition. These include the source of saliva, flow rate, degree of hydration of the subject, nature of the stimulus, duration of stimulation, plasma composition, the time of day at which samples are collected, and the serial dependency of saliva samples (the effect of previous stimulation on the composition of saliva collected subsequently). The importance of some of these factors has only recently been appreciated. In addition, the experimental design of previous studies rarely included a group of control subjects kept on a constant diet. The effect of nutrition on saliva was reviewed briefly by Krasse.1

Digestive Enzymes of the American Lobster (Homarus americanus)

Abstract: Gastric juice of lobster hydrolyzed the following substrates: triolein (enzymic activity: lipase), tributyrin ("lipase"), Azocoll(R) (proteinase), TAME and BAEE ("trypsin"), ATEE ("chymotrypsin"), HPLA ("carboxypeptidase A"), DNA (deoxyribonuclease), RNA (ribonuclease), p-nitrophenyl-phosphate (phosphatase), and p-nitrophenyl-N-acetyl-β-glucosaminide (chitobiase). (The quotation marks signify that the substrate specificities are typical of the quoted enzymes of mammalian or microbial origin. The spectra of specificities, however, of these enzymes and the corresponding enzymes of lobster are not necessarily identical.) Very low activities were found against RBB-starch (amylase) p-nitro-phenyl-α-(and β)-glucopyranoside(α- and β-glucosidase), p-nitrophenyl-β-galactopyranoside (β-galactosidase), and chitin azure (chitinase). No activities corresponding to phospholipase (lecithin), carboxypeptidase B (benzoylglycyl-L-arginine), elastase, glycylglycine dipeptidase, or leucine aminopeptidase were found.Gel filtr...

The effect of trypsin inhibitors on pancreatopeptidase E, trypsin, chymotrypsin and amylase in the pancreas and intestinal tract of chicks receiving raw and heated soya-bean diets.

Abstract: 1. Feeding on a raw soya-bean diet (RSD) increased the levels of trypsin, chymotrypsin and pancreatopeptidase E but decreased the level of amylase in the pancreas of chicks as compared to a heated soya-bean diet (HSD), while supplementation of HSD with soya-bean trypsin inhibitors increased the activity of all four enzymes. HSD + trypsin inhibitors caused significant enlargement of the pancreas but only a slight depression in growth rate.2. Fasting for 24 h of chicks previously given RSD and HSD increased the activity of all four enzymes but the increase was much greater in chicks previously given RSD than in those previously given HSD.3. Feeding RSD for 4 d to chicks previously adapted to HSD resulted in a dramatic inhibition in growth rate, a small increase in pancreas weight, and an increase in the activity of all proteolytic enzymes, while no change in the amylase was detectable.4. Trypsin, chymotrypsin and pancreatopeptidase E activities were assayed in the contents of the small intestine and caecum of chicks fed on RSD or HSD over a period of 35 d. Trypsin and chymotrypsin activities in the small intestine were lower in chicks fed on RSD while pancreatopeptidase E activity was almost equal or even higher in RSD-fed chicks, especially at the age of 35 d. Trypsin activity in the caecum of RSD-fed chicks was lower at all stages of the experiment, while the pancreatopeptidase E and chymotrypsin activities in the caecum of RSD-fed chicks exceeded the levels in the HSD group at the age of 21 and 35 d respectively. It would appear therefore that pancreatopeptidase E may play an important part in overcoming the inhibition of the proteolytic activity in the intestine of chicks fed on RSD.

Production of amylase in liquid culture by a strain of Aspergillus oryzae.

Abstract: The effect of different media and pH on the formation of amylase by Aspergillus oryzae EI 212 is described. Depending upon the composition of the medium and growth conditions, the fungus was found to secrete α- or β-amylase, or both. Some of the properties of the partially purified α-amylase were found to be different from α-amylases from other sources.

Properties of the Amylase from Halobacterium halobium

Abstract: Halobacterium halobium amylase had optimal activity at pH 6.4 to 6.6 in sodium β-glycerophosphate buffer containing 0.05% NaCl at 55 C; Ca2+ was not required. End products from amylose were maltose, maltotriose, and glucose. The amylase, which was devoid of transglucosylase activity, had a multichain attack mechanism.

Enzymes of starch metabolism in the developing rice grain.

Abstract: The levels of starch, soluble sugars, protein, and enzymes involved in starch metabolism-alpha-amylase, beta-amylase, phosphorylase, Q-enzyme, R-enzyme, and starch synthetase -were assayed in dehulled developing rice grains (Oryzasativa L., variety IR8). Phosphorylase, Q-enzyme, and R-enzyme had peak activities 10 days after flowering, whereas alpha- and beta-amylases had maximal activities 14 days after flowering. Starch synthetase bound to the starch granule increased in activity up to 21 days after flowering. These enzymes (except the starch synthetases) were also detected by polyacrylamide gel electrophoresis. Their activity in grains at the midmilky stage (8-10 days after flowering) was determined in five pairs of lines with low and high amylose content from different crosses. The samples had similar levels of amylases, phosphorylase, R-enzyme, and Q-enzyme. The samples consistently differed in their levels of starch synthetase bound to the starch granule, which was proportional to amylose content. Granule-bound starch synthetase may be responsible for the integrity of amylose in the developing starch granule.

New Chromogenic Substrate for Determination of Serum Amylase Activity

Abstract: A rapid assay for serum amylase activity has been developed based on the use of a new chromogenic substrate—Cibachron Blue—amylose. The procedure requires 0.1 ml of serum and measures the production of soluble chromogen, the formation of which is linear with enzyme activity. Normal human serum has a mean amylase activity of 118 ± 37 (SD) mg dye/100 ml/15 min or, in international units, 78 ± 24 (SD) µg dye/min/ml. Excellent correlation was observed with a saccharometric amylase procedure.

Inhibition of exocrine pancreatic secretion by glucagon and D-glucose given intravenously.

Abstract: The actions of glucagon and D-glucose on blood glucose and exocrine pancreatic secretion in response to secretin were studied in unanesthetized dogs with chronic pancreatic fistulas, gastric fistulas, and a gastroenterostomy which diverted gastric acid from the duodenum. Both glucagon and D-glucose, when given intravenously, produced significant and dose-related inhibition of the volume of pancreatic secretion and of the protein, amylase, lipase, and protease outputs. There was a linear inverse relationship between pancreatic enzyme output and blood glucose levels following glucagon or D-glucose infusion. Statistical analysis of the data showed that the slopes of the lines relating blood glucose to amylase, to lipase, and to protease were significantly different from each other, indicating preferential inhibition for amylase.

A study of sieve element starch using sequential enzymatic digestion and electron microscopy

Abstract: The fine structure of plastids and their starch deposits in differentiating sieve elements was studied in bean (Phaseolus vulgaris L) Ultrastructural cytochemistry employing two carbohydrases specific for different linkages was then used to compare the chemical nature of "sieve tube starch" (the starch deposited in sieve elements) with that of the ordinary starch of other cell types Hypocotyl tissue from seedlings was fixed in glutaraldehyde, postfixed in osmium tetroxide, and embedded in Epon-Araldite Treatment of thin sections on uncoated copper grids with α-amylase or diastase at pH 68 to cleave α-(1 → 4) bonds resulted in digestion of ordinary starch grains but not sieve element grains, as determined by electron microscopy Since α-(1 → 6) branch points in amylopectin-type starches make the adjacent α-(1 → 4) linkages somewhat resistant to hydrolysis by α-amylase, other sections mounted on bare copper or gold grids were treated with pullulanase (a bacterial α-[1 → 6] glucosidase) prior to digestion with diastase Pullulanase did not digest sieve element starch, but rendered the starch digestible subsequently by α-amylase Diastase followed by pullulanase did not result in digestion The results provide evidence that sieve element starch is composed of highly branched molecules with numerous α-(1 → 6) linkages

Enzymic Mechanism of Starch Breakdown in Germinating Rice Seeds: III. alpha-Amylase Isozymes.

Abstract: The formation of amylase isozymes in germinating rice (Oryza sativa) seeds was studied by isoelectric focusing on polyacrylamide gel disc electrophoresis. Time sequence comparisons of the amylase zymogram were made between extracts from gibberellic acid-treated embryoless and embryo-attached half-endosperm of rice seeds. In both cases, 4 major and 9 to 10 minor isozyme bands were detectable at the maximal stage of the enzyme induction. However, in the embryo-attached half-seeds, bands started to diminish after the 5th day of incubation, in agreement with the results of time sequence analyses of enzyme activities. Nearly identical patterns of amylase isozyme bands on a polyacrylamide gel disc electrophoresis in combination with isoelectric focusing indicate the intrinsic role of gibberellic acid in the starch breakdown in germinating rice seeds. We tentatively assign the newly synthesized enzymes to be alpha-amylases based on experimental results concerning the lability of the preparation on a prolonged treatment at pH 3.3 and the stability on heat treatment for 15 minutes at 70 C.

Usefulness of Serum Lipase, Esterase, and Amylase Estimation in the Diagnosis of Pancreatitis—a Comparison

Abstract: Sera of 19 individuals with clinically established pancreatitis were analyzed for lipase activity on emulsion-type and aqueous substrates. Serum amylase concentrations were compared. Results obtained on the first and subsequent days of hospitalization clearly indicated that methods for lipase determination in which an emulsion-type substrate is used are of the greatest aid in diagnosing pancreatitis (90 to 92% of patients with pancreatitis had supranormal results). Amylase determinations were nearly as useful as an index (78% correlation), but "lipase" values obtained with methods in which aqueous substrates are used had limited clinical usefulness (29 and 32% correlation). Serum lipase elevations in cases of pancreatitis were generally greater than amylase elevations, with some exceptions. Serum lipase should be determined with a method in which emulsion-type substrates are used. Lipase and amylase values supplement one another.

The interrelationships of pancreatic enzymes in human duodenal aspirate

Abstract: The interrelationships of proteolytic enzymes and amylase have been studied in the duodenal aspirate obtained from subjects with normal and abnormal pancreatic function during stimulation with secretin and pancreozymin. While the relationship of trypsin to chymotrypsin was independent of stimulus and presence of pancreatic disease the ratio of proteolytic enzymes to amylase rose when the degree of stimulation of the pancreas was increased. Patients with recent acute pancreatitis and with chronic pancreatitis tended to have more severe impairment of secretion of proteolytic enzymes than of amylase. In routine tests of pancreatic function both proteolytic and non-proteolytic enzymes should be measured, both because an abnormal ratio may be of diagnostic significance and because the two different groups of enzymes provide mutual checks of the secretory capacity of pancreatic enzymes.

New Amylase Substrate and Assay Procedure

Abstract: This report describes a new procedure for serum amylase assay. A novel substrate, dyed amylopectin, is used that combines the advantages of saccharogenic and amyloclastic methods. Serum amylase hydrolyzes the dyed amylopectin into ethanol-soluble fragments, which are quantified colorimetrically after serum proteins and unhydrolyzed substrate are precipitated with alcoholic tannic acid. The substrate is prepared by coupling Reactone Red 2B to amylopectin in alkaline solution. Unreacted dye is removed by gel filtration. The clear red solution of dyed amylopectin is buffered and diluted to a standard concentration; it can be preserved indefinitely by lyophilization. The assay procedure is the following: 0.2 ml serum is added to 1 ml substrate at 37°C. After incubation for 10 min, 5 ml of alcoholic tannic acid are added, and the mixture is centrifuged. Absorbance of the supernatant solution at 540 nm is a linear function of amylase activity. Serum blanks are not required. The results correlate well with the saccharogenic assay of Somogyi.

Purification and characterization of the amylase of B. subtilis NRRL B3411

Abstract: The amylase of Bacillus subtilis NRRL B3411 has been purified and partially characterized. The specific activity can be increased from 300,000 units/g to 6,000,000 units/g with a 60% recovery of total units. The purified material consists of one major and one trace anodic component as determined by disc gel electrophoresis. The molecular weight was 48,000 as determined by bio-gel filtration; the molecular weight was 44,900 ± 2400 as determined by sedimentation equilibrium methods. This purified enzyme is stable at, 70°C in the presence of 0.01 M Ca++ and 0.1 M NaCl over a broad pH range from 5.5–9.5. The pH activity profile indicates optimum activity at pH 6.0. This amylase exhibits maximum activity at 60°C. The enzyme is a liquefying α-amylase as determined by analysis of hydrolysis products and immunological studies.

Sugar changes during the preparation of burukutu beer

Abstract: In the study of sugar changes during preparation of Burukutu beer, samples analysed were obtained from a local brewer. A freshly prepared drink was analysed together with gari – a starch adjunct. The pattern of sugar changes during this 5-day malting stage was also followed. Twelve sugars were identified in the beverage while six sugars were shown to be present in the gari adjunct. Analyses of both germinated and ungerminated sorghum grains show that no new types of sugars resulted from the process of germination. In the study of amylase activity of the malts it was shown that detectable amylase activity began from the second day of germination onwards. Because of the relatively high concentrations of maltooligosaccharides in the drink, starch amylosis should have continued during the fermentation stage.

α-amylase-isozymes in gibberellic-acid-treated barley half seeds.

Abstract: The presence of multiple forms of α-amylase in gibberellic acid-treated embryoless barley half-seeds was demonstrated by separation on diethylaminoethyl-Sephadex and isoelectric focusing polyacrylamide gel disc electrophoresis. Two major α-amylase fractions (A and B), each consisting of two to three isozyme components, were purified. α-Amylase fractions A and B were distinguishable in their reaction patterns. The optimal pH of fraction A α-amylase was found to reside in the acidic side (pH 5.0), as was determined by analyzing the reducing sugars formed as well as the paper chromatographic detection of reaction products. At neutral pH, 6.9, fraction A exhibited weak amylolytic activity in forming maltose. The α-amylase activity in fraction A was markedly stimulated by heat treatment (70 C/15 minutes). Fraction B, constituting a major part of amylases in the endosperm extract, was also found to be composed of α-amylase, as evidenced by the loss of enzyme activity upon allowing fractions A and B to stand at pH 3.3 for a prolonged period. The possible physiological function of the two different types of α-amylase in the carbohydrate breakdown of barley seeds is discussed.

Electrophoretic studies of α‐amylase in wheat. II

Abstract: The lack of correlation between the degree of sprouting and the α-amylase activity in wheat and rye, as well as the apparent variation in the falling number during ripening, can be explained as the result of two amylase systems acting during different stages of the development of the grain. During the early stages of the development of the grain, α-amylase is continuously inactivated. This process is reversible, however, and when the evaporation of moisture is retarded the α-amylase activity increases as a consequence of the higher amount of dissolved enzyme. This results in an occasional decrease in the falling number, which might amount to more than 50 sec. During germination, a new kind of α-amylase develops. The synthesis of the new form of α-amylase is irreversible and causes a much greater and permanent reduction in the falling number. During the initial stages of germination, the amylase activity thus increases owing to the combined action of both the original amylase and the new form. The two kinds of amylases show different electrophoretic patterns. Drying the grain after harvesting reduces the activity of ‘green’ amylase (amylase in unripened grain), which might explain the frequent observations of increasing falling number during storage.

An improved amylase assay using a new starch derivative.

Abstract: A method for the assay of α-amylase using Remazolbrilliant blue starch (RBB-starch), an insoluble, blue starch derivative, is described. A suspension of the starch is solubilized by the hydrolytic action of amylase, and after acidification to stop enzymatic activity and filtration to remove remaining insoluble RBB-starch, the amount of solubilized color can be related spectrophotometrically to the enzyme concentration. A kinetically valid procedure applicable to the routine clinical laboratory is presented. Results obtained with this new method are compared statistically with results of representative amyloclastic and saccharogenic technics.

The Characterization of Starch and its Components. Part 2. The Semi-Micro Estimation of the Starch-Content of Cereal Grains and Related Materials†

Abstract: A rapid, semi-micro method has been developed for the estimation of the amount of starch in cereal grains. The method is suitable for samples of flour in the range 7.5–20 mg, and hence the amount of starch in a single grain can be readily determined. The procedure – which is given in detail – involves first solubilizing the starch by hot aqueous calcium chloride, and then subjecting the extract to the concurrent action of a-amylase and amyloglucosidase. A quantitative determination of the resultant glucose is then made by the use of glucose oxidase. Relatively few manipulations are involved. This technique is applicable to a wide range of cereals, including the high-amylose-content, genetic mutants of maize. The method has a reproduceability of ± 1.5 %. The new procedure has been compared with methods based on polarimetry and perchloric acid extraction.