OBJECTIVETo compare the effect of citric acid stimulation on salivary alpha-amylase (sAA), total protein (TP), salivary flow rate, and pH value between Pi deficiency (PD) children and healthy children, thereby providing evidence for Pi controlling saliva theory.

METHODSTwenty PD children were recruited, and 29 healthy children were also recruited at the same time. Saliva samples from all subjects were collected before and after citric acid stimulation. The sAA activity and amount, TP contents, salivary flow rate, and pH value were determined and compared.

RESULTS(1) Citric acid stimulation was able to significantly increase salivary flow rate, pH value, sAA activities, sAA specific activity and sAA amount (including glycosylated and non-glycosylated sAA amount) in healthy children (P<0.05), while it could markedly increase salivary flow rate, pH value, and glycosylated sAA levels in PD children (P<0.05); (2) Although there was no statistical difference in determined salivary indices between the two groups (P>0.05), salivary indices except salivary flow rate and glycosylated sAA levels decreased more in PD children. There was statistical difference in sAA activity ratio, sAA specific activity ratio, and the ratio of glycosylated sAA levels between PD children and healthy children (P<0.05).

CONCLUSIONPD children had decreased response to citric acid stimulation.

Child ; Citric Acid ; therapeutic use ; Humans ; Medicine, Chinese Traditional ; Saliva ; Salivary alpha-Amylases ; metabolism ; alpha-Amylases

OBJECTIVETo optimize different processing techniques of Massa Medicata Fermentata.

METHODSingle factor test was adopted, with the amylase activity of Massa Medicata Fermentata as the assessment indicator, to observe the influence of such factors as fermentation time and mixture techniques of active pharmaceutical ingredients on the amylase activity of Massa Medicata Fermentata. Meanwhile, Massa Medicata Fermentata prepared with the optimum processing techniques and superior and inferior products of Massa Medicata Fermentata in the market were compared in amylase activity, soluble starch content and soluble polysaccharide content.

RESULTThe optimum fermentation time was 7 days. Adzuki bean shall be boiled before mixed with other materials. Artemisia annua, Polygonum hydropiper and Cocklebur grass shall be evenly mixed water decoction. The amylase activity, the soluble starch content and the soluble polysaccharide content of fermented Massa Medicata Fermentata could reach to 49.372 mg x min(-1) x g(-1), 7.967%, and 16.65% respectively, significantly higher than the two types of Massa Medicata Fermentata sold in the market.

CONCLUSIONAccording to the optimum processing techniques, Adzuki beans were smashed and mixed equally with flour and Armeniacae Semen Amarum powder, and then successively added with A. annua, P. hydropiper and C. grass for even mixture. The fermentation time was 7 days.

Amylases ; metabolism ; Drug Compounding ; Fermentation ; Medicine, Chinese Traditional

Studies on the variation of amylase, lipase and lrotease activity of Whitmania pigra in 0-6 months old using 3, 5-dinitro- salicylic acid colorimetry, right-nitrophenyl palmitate ester (ρ-NPP) colorimetry and folin-phenol method. The results showed that pro- tease activity remained low before 1.5 months old and with the highest activity in 2 months old, but after showing a small peak in 4 months, alkaline protease rapid declined. Amylase was low at born, then gradually increased the activity of the highest in 2.5 months old. Lipase with a strong vitality at birth, then 1 month with minimum and 2 months peaked, but appeared a small peak in 4 months old. In summary, only lipase exhibits strong activity at birth, lipase with the strongest activity in the digestive tract during develop- ment. Protease, lipase and amylase with the strongest activity at 2-3 months old, but were decreased after 4 months old.
Age Factors ; Amylases ; metabolism ; Animals ; Leeches ; enzymology ; Lipase ; metabolism ; Peptide Hydrolases ; metabolism

OBJECTIVETo extract polysaccharides from Dioscorea opposita by alpha-amylase and ultrasound.

METHODThe optimum condition of enzyme-assisted extraction has been obtained through orthogonal test according to the yield of polysaccharides.

RESULTA higher yield of polysaccharides was achieved at 55 degrees C, pH 5.5 , with a load of alpha-amylase 10 mg for 1.0 hour, the extraction rate was increased by in compare of sonolysis treatment alone.

CONCLUSIONTo carry out an enzyme treatment before ultrasound--assisted extraction elevated the yield of polysaccharides.

Dioscorea ; chemistry ; Hydrogen-Ion Concentration ; Polysaccharides ; chemistry ; isolation & purification ; metabolism ; alpha-Amylases ; metabolism

Synaptotagmin is a Ca2+ sensing protein, which triggers a fusion of synaptic vesicles in neuronal transmission. Little is known regarding the expression of Ca2+ - dependent synaptotagmin isoforms and their contribution to the release of secretory vesicles in mouse and rat parotid acinar cells. We investigated a type of Ca2+ - dependent synaptotagmin and Ca2+ signaling in both rat and mouse parotid acinar cells using RT-PCR, microfluorometry, and amylase assay. Mouse parotid acinar cells exhibited much more sensitive amylase release in response to muscarinic stimulation than did rat parotid acinar cells. However, transient [Ca2+]i increases and Ca2+ influx in response to muscarinic stimulation in both cells were identical, suggesting that the expression or activity of the Ca2+ sensing proteins is different. Seven Ca2+ - dependent synaptotagmins, from 1 to 7, were expressed in the mouse parotid acinar cells. However, in the rat parotid acinar cells, only synaptotagmins 1, 3, 4 and 7 were expressed. These results indicate that the expression of Ca2+ - dependent synaptotagmins may contribute to the release of secretory vesicles in parotid acinar cells.
Synaptotagmins/*metabolism ; Signal Transduction ; Rats ; Protein Isoforms/metabolism ; Parotid Gland/cytology/*metabolism ; Muscarinic Agonists/pharmacology ; Mice ; Exocytosis/drug effects/physiology ; Carbachol/pharmacology ; Calcium/metabolism/*physiology ; Animals ; Amylases/secretion

The alpha-amylase (EC 3.2.1.1) from the Gram-positive Alicyclobacillus acidocaldarius was one kind of thermoacidophilic enzyme, with optimal temperature and pH of 75 degrees C and 3, respectively. The nucleotide sequence of the gene amy was cloned by PCR. The gene amy was 3901bp long, comprising one open reading frame encoding a polypeptide of 1301 amino acids. The calculated molecular weight of the alpha-amylase AMY was about 140kD. The gene amy was expressed in E. coli BL21 (DE3) and Pichia pastoris respectively, and both of the cloned proteins had bioactivity. The activity of amylase expressed in P. pastoris was further testified by amylase activity staining. The alpha-amylase expressed in P. pastoris had been purified and characterized. The apparent molecular weight of that was about 160kD according to SDS-PAGE. The optimum of pH for the enzyme was pH 3.2 as the native enzyme was; but the optimum of temperature was 65 degrees C and a little lower than that of the native enzyme. Above 50% of relative activity remained after incubation for 30 minutes in 70 degrees C. So the enzyme expressed by P. pastoris was also thermoacidophilic. Moreover some sequence was cloned by PCR, which ranged from + 1174 bp to + 3288 bp in the gene amy, encoding 705 amino acids with the calculated molecular weight of 79kD. The truncated gene amy' was expressed in E. coli BL21 (DE3) induced by 1 mmol/L IPTG, and the expressed enzyme also retained alpha-amylase activity.
Bacillus ; enzymology ; genetics ; Bacterial Proteins ; genetics ; isolation & purification ; metabolism ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Pichia ; genetics ; metabolism ; alpha-Amylases ; genetics ; isolation & purification ; metabolism

OBJECTIVEIn order to explore the toxic effects of cadmium on functions of endocrine and exocrine of pancreas.

METHODS96 SD rats were administered with cadmium at different doses (0, 50, 100, 200 mg/L) by drinking water for 30 days, 60 days and 90 days. The contents of cadmium and zinc in the blood and pancreas, also the glucose level in blood and urine, the levels of insulin and the activity of amylase were determined. The gene expression of metallothionein (MT), insulin and pancreatic amylase were also measured.

RESULTSThe results showed that the contents of cadmium in the serum and pancreas were higher than that of the control groups (P < 0.05). The contents of zinc in serum were decreased in the groups of 100 and 200 mg/L cadmium at the 90-day. As well as increased zinc in pancreas. The gene expression of insulin was not different compared with those of the control group except the middle-dose group at the 60-day. And the expression of amylase were higher in the groups of 100 and 200 mg/L cadmium at the 60-day and the 90-day. The expression of MT-1 and -2 were higher in the pancreas after cadmium administration.

CONCLUSIONIt is suggested that cadmium could be accumulated in the pancreas and caused the change of the zinc levels. Then it resulted in the change of the expression of gene and protein, and influence of the functions of both endocrine and exocrine in pancreas.

Amylases ; metabolism ; Animals ; Cadmium ; toxicity ; Female ; Insulin ; metabolism ; Male ; Metallothionein ; metabolism ; Pancreas ; drug effects ; metabolism ; Rats ; Rats, Sprague-Dawley ; Zinc ; blood

Genetic modification of barley variety can be an efficient way to improve beer quality. The objective of this study was to understand the effect of trxS gene on hydrolases activities in transgenic and non-transgenic barley seeds. The results showed that alpha-amylase, free beta-amylase and limit dextrinase activity were increased in transgenic seeds in comparison with non-transgenic seeds. Sulfhydryl content of protein in transgenic seeds was also higher than that in non-transgenic seeds, suggesting that trxS gene could express in barley seeds, which opens a new way for breeding new barley varieties to improve beer quality.
Germination ; genetics ; Glucosyltransferases ; metabolism ; Hordeum ; enzymology ; genetics ; Plants, Genetically Modified ; enzymology ; genetics ; Seeds ; enzymology ; genetics ; Sulfhydryl Compounds ; metabolism ; Thioredoxins ; genetics ; alpha-Amylases ; metabolism ; beta-Amylase ; metabolism

Supplement effects of ions, sugars, and amino acids on the thermostability of liquefying type alpha-amylase from Bacillus subtilis were examined. The addition of 1 mmol/L Ca2+ or about 50 mmol/L Na+ remarkably stimulated the thermostability of this enzyme among ions examined. The thermostability of the enzyme was enhanced and reduced by the extrinsic addition of 50 mmol/L acidic amino acid such as glutamic acid and alkaline amino acid of the concentrations of sugars from 0 to 1000 mmol/L the thermostability of alpha-amylase increased almost such as arginine, respectively. With the increases linearly. By the co-existence of Na+ or K+ with some amino acids or sugars the thermostability of this enzyme was fairly increased. The changes in the fluorescence intensity of alpha-amylase were examined as a function of the incubation temperature on the enzyme, which showed a good agreement with those of residual activities.
Amino Acids ; pharmacology ; Calcium ; pharmacology ; Carbohydrates ; pharmacology ; Enzyme Stability ; Protein Conformation ; Sodium ; pharmacology ; Temperature ; alpha-Amylases ; chemistry ; metabolism

Thermostable alpha-amylase from Pyrococcus furious is an important industrial enzyme in brewing and alcohol production. Eexpression of the thermostable a-amylase in plants can reduce greatly costs in the production of alcohol using crop plants. A chloroplast expression vector, p64A, containing the thermostable alpha-amylase gene from Pyrococcus furious, was constructed with clpP-trnL-petB-chlL-rp123-rpl2 as Chlamydomonas reinhardtii plastid homologous recombinant fragments and spetinomycin-resistant aadA gene as select marker. The plasmid p64A was transferred into the chloroplast genome of C. reinhardtii by the biolistic method. Nine independently transformed lines were obtained by 100 mg/L spectinomycin selection. PCR amplification, Southern blot analysis of the transgene and cultivation in the dark all showed that the a-amylase gene had been integrated into chloroplast genome of C. reinhardtii. The activity of amylase expressed in the chloroplast of C. reinhardtii was detected by amylase activity assay and found to be as high as 77.5 u/g fresh weight of cells. These experimental results demonstrated the possibility of using transgenic chloroplasts of plant as bioreactors for production of industrial enzymes.
Chlamydomonas reinhardtii ; genetics ; Chloroplasts ; genetics ; Enzyme Stability ; Plasmids ; Polymerase Chain Reaction ; Pyrococcus furiosus ; enzymology ; alpha-Amylases ; chemistry ; genetics ; metabolism