Effects of Exogenous Amylases and Metal Ions on the Amylase Specific Activities and Starch Degradation of the Upper Leaves of ‘KRK_(26)' during Flue-curing

Objective] The aim of this study was to investigate the effects of exoge-nous amylases and Ca2＋, Mn2＋ and K＋ on the amylase specific activities and starch degradation of the upper leaves of ＇KRK26＇ planted in Yunnan Province during flue-curing. [Method] The amylase specific activities and starch degradation of the leaves were determined by using spectrophotometry. [Result] The 8 U/g exogenous α-amy-lase could improve the specific activity of the leaf α-amylase at yel owing and color-fixing stages, but could not at stem-drying stage, and similarly, the 80 U/g exoge-nous β-amylase could improved the specific activity of the leaf β-amylase at the yel owing stage and the early period of color-fixing stage. The leaf starch could be enhanced to degrade by the exogenous α- or β-amylases and the enhancing effect of the former was stronger than that of the later. 1.50 mg/ml Ca2＋ improved the specific activity of the leaf （α＋β）-amylase mainly due to its enhancing effect on the leaf α-amylase, and increased the starch degradation. 4 mmol/L Mn2＋ inhibited the leaf α-amylase from yel owing to the early period of color-fixing and the β- and （α＋β）-amylases from the yel owing to the later period of color-fixing, but enhanced the leafα-amylase from the later period of color-fixing to the later period of stem-drying and the β- and （α＋β）-amylases at the later period of stem-drying. Meanwhile, Mn2＋ ham-pered the starch degradation during yel owing, but promoted it from the early period of color-fixing to stem-drying. 1 mg/ml K＋ enhanced the leaf α-, β- and （α＋β）-amy-lases during the yel owing stage, but lowered them from the early period of color-fix-ing to the later period of stem-drying, and always inhibited the leaf starch degrada-tion. [Conclusion] The exogenous α-, β- amylases and Ca2＋ of suitable concentra-tions could be used to treat the tobacco leaves before flue-curing to improve the leaf starch degradation during the curing.

Analysis of Amylase and Superoxide Dismutase Isozymes During the Germination Process of Emmenopterys henryi Oliv Seeds

[ Objective] The study was to understand the changes of amylase（AMY） and superoxide dismutase（SOD） isozymes during the ger- mination process of Emmenopterys henryi Oliv seeds. [ Metbod] By employing polyacrylamide gel electrophoresis method, the expressions of AMY and SOD isozymes during seed germination process were analyzed. ~ Result] The main AMY bands remained strong during the whole peri- od and a new band A2 appeared in the middle and late period of seed germination. Some new SOD bands occurred at the early stage, then be- came weak or disappeared in the middle period, and band S6 became intense in the late peried. [ Conclusion.] The expression of AMY and SOD isozyme gene has temporal difference during germination of E. henryi Oliv seeds.

Screening and Identification of Amylase-producing Strain from Arctic Ocean and Optimization of Enzyme Producing Conditions

[Objective] The aims were to investigate the screening and identification of amylase-producing marine bacteria from Arctic sea and the optimization of the amylase producing conditions. [Method] A high-yield strain for producing amylase named ArcB84A was isolated from a total of 156 marine bacteria of Arctic sea. Then,the morphological identification of the strain,molecular identification of 16S rRNA and optimization of fermentation conditions were conducted. [Result] ArcB84A strain was a member of Pseudoalteromonas genus. The optimum conditions for enzyme production of B84A strain included that,the initial pH value of the medium was 7.0-8.0,and the best carbon and nitrogen sources respectively were 5‰ glucose and peptone. Surfactants including TritonX-100,Tween20 and Tween80 could increase amylase activity of the strain,in which,the effect of 10‰ Tween80 was the most obvious.

Lipase and pancreatic amylase activities in diagnosis of acute pancreatitis in patients with hyperamylasemia

BACKGROUND: Measurement of total serum amylase (AMY) is the most widely used biochemical test for the diagnosis of acute pancreatitis, but it is commonly considered a nonspecific marker. To improve the biochemical diagnosis of acute pancreatitis, lipase ( LIP ) and pancreatic amylase (PAMY) have been tested in recent years. The present study was designed to evaluate whether serum LIP and pancreatic PAMY tests could replace total amylase test to improve diagnostic efficiency in the evaluation of acute pancreatitis in patients with hyperamylasemia. METHODS: LIP and PAMY values were determined in serum samples from 92 patients with hyperamylasemia. Reference values for each enzyme were derived from serum samples of 147 healthy subjects. The activities of LIP and PAMY in patients with various diseases were shown directly by the boxplot graph. The diagnostic accuracy of LIP and PAMY was defined as the area under the receiver operating characteristic (ROC) curve. Their sensitivity and specificity in detecting acute pancreatitis at varying cutoff points were shown by the curve, and the best cutoff value for each enzyme was shown by the modified ROC curve. The diagnostic values of LIP, PAMY and LIP + AMY with each upper limit of reference range (ULR) were compared with the corresponding best cutoff values. RESULTS: The references values of LIP and PAMY were 12.2-47.6 U/L and 28-95 U/L, respectively. These values in patients with acute pancreatitis were higher than those patients with other diseases. The areas under the ROC curve ( AUC) of LIP and PAMY were 0. 799 and 0. 792, respectively, With the best diagnostic cutoff point of maximum (sensitivity + specificity) -100%, we obtained values of 97.9 U/L(LIP97.9 =2. 06 × ULR) for LIP and 209 U/L (PAMY209 =2.20 ×ULR) for PAMY. The best cutoff values for LIP, PAMY and LIP +AMY demonstrated the specificity, positive predictive value, and diagnostic efficiency higher than the corresponding ULRs. CONCLUSIONS: Serum LIP and PAMY are specific for the pancreas and might replace total amylase for the diagnosis of acute pancreatitis in hyperamylasemia patients. LIP97.9 is more efficient than PAMY209 in the diagnosis of acute pancreatitis. A combined test of both enzymes is not superior to single test of either enzyme in diagnostic accuracy.

Relationship between post-ERCP pancreatitis and the change of serum amylase level after the procedure

AIM: To clarify the relationship between the change of serum amylase level and post-ERCP pancreatitis. METHODS: Between January 1999 and December 2002, 1291 ERCP-related procedures were performed. Serum amylase concentrations were measured before the procedure and 3, 6, and 24 h afterward. The frequency and severity of post-ERCP pancreatitis and the relationship between these phenomena and the change in amylase level were estimated. RESULTS: Post-ERCP pancreatitis occurred in 47 patients (3.6%). Pancreatitis occurred in 1% of patients with normal amylase levels 3 h after ERCP, and in 1%, 5%, 20%, 31% and 39% of patients with amylase levels elevated 1-2 times, 2-3 times, 3-5 times, 5-10 times and over 10 times the upper normal limit at 3 h after ERCP, respectively (level < 2 times vs ≥ 2 times, P < 0.001). Of the 143 patients with levels higher than the normal limit at 3 h after ERCP followed by elevation at 6 h, pancreatitis occurred in 26%. In contrast, pancreatitis occurred in 9% of 45 patients with a level higher than two times the normal limit at 3 h after ERCP followed by a decrease at 6 h (26% vs 9%, P < 0.05). CONCLUSION: Post-ERCP pancreatitis is frequently associated with an increase in serum amylase level greater than twice the normal limit at 3 h after ERCP with an elevation at 6 h. A decrease in amylase level at 6 h after ERCP suggests the unlikelihood of development of post-ERCP pancreatitis.

Drain amylase value as an early predictor of pancreatic fistula after cephalic duodenopancreatectomy

AIM: To determine predictors of clinically relevant pancreatic fistulas (CRPF) by measuring drain fluid amylase (DFA) in the early postoperative period.

MICROSTRUCTURE AND AMYLASE ACTIVITY OF CHINESE CABBAGE SEED DURING DRYING PROCESS

A seed is the carrier of life,and research on the heat and mass transfer of a seed has already stridden toward the level of microcosm,such as cell protoplasm,cell organism,and molecular membrane.By means of a transmission electron microscope,the authors of this paper observed the microstructure of cotyledon tissue slices of the Chinese cabbage seed with a moisture content of 13% (on dry basis) and that with a moisture content of 4.3% (on dry basis) for drying 2 h at 45 ℃.The compared result was that only wrinkles had been discovered on the cell walls of the seed dried for 2 h,without any significant change for other organelles.Study on the enzyme activity shows that after a germination for 48 h,the relative activity of α amylase of the Chinese cabbage seed dried for 2 h at 45 ℃,decreased by 5.8%,whereas that of the seed dried 2 h at a temperature of 67 ℃ decreased by 30.1%.This work shows that the drying factors have greatly influence on the seed microstructure,enzyme activity,which is directly positive to seed viability.Combined with the analysis of the critical safe drying temperature of the vegetable seed,it can be concluded that enzyme activity is also the function of the drying temperature,the moisture content and the drying time.

Effects of Mercury Stress on Wheat Growth,POD and Amylase Activity

[Objective] The aim was to study the effect of mercury stress on seed germination and seedlings growth.[Method]using Zhengzhou 9023 as the experimental material and cultured in water,to study the effect of the germinating rate,seedling height,root length,seedling＇s fresh weight,the activities of peroxidase（POD）and amylase in leaf,root and germinating embryo at different concentrations of Hg2＋（0.025,0.050,0.100,0.200,0.300,0.400,0.500 mmol/L）.[Result]Low concentrations of Hg2＋（≤ 0.10 mmol/L）have little effect on seed germination,seedling height,root length and fresh weight;high concentrations of Hg2＋（ 0.10 mmol/L）have significantly inhibited seed germination and seedling growth.Low concentration of Hg2＋（≥ 0.025 mmol/L）could increase POD activity and inhibit the amylase activity significantly,and the effects have increased with the increasing of Hg2＋ concentrations.[Conclusion]Hg2＋ stress could change the activities of POD and amylase in leaf,root and germinating embryo,influence the energy and substrate supply which was required for normal metabolism of lipid oxidation,and inhibit seedling growth ultimately.

Monitoring salivary amylase activity is useful for providing timely analgesia under sedation

AIM:To detect the criteria and cause of elevated salivary amylase activity(sAMY)in patients undergoing endoscopic submucosal dissection(ESD)under sedation.METHODS:A total of 41 patients with early gastric cancer removed via ESD under deep sedation(DS)were enrolled.The perioperative sAMY,which was shown as sympathetic excitements(SE),was measured.The time at which a patient exhibited a relatively increased rate of sAMY compared with the preoperative baseline level(IR,%)≥100%(twice the actual value)was assumed as the moment when the patient received SE.Among the 41 patients,we focused on 14 patients who exhibited an IR≥100% at any time that was associated with sAMY elevation during ESD(H-group)and examined whether any particular endoscopic procedures can cause SE by simultaneously monitoring the sAMY level.If a patient demonstrated an elevated sAMY level above twice the baseline level,the endoscopic procedure was immediately stopped.In the impossible case of discontinuance,analgesic medicines were administered.This study was performed prospectively.RESULTS:A total of 26 episodes of sAMY eruption were considered moments of SE in the H-group.The baseline level of sAMY significantly increased in association with an IR of>100% at 5 min,with a significant difference(IR immediately before elevation/IR at elevation of sAMY=8.72±173/958±1391%,P<0.001).However,effective intervention decreased the elevated sAMY level immediately within only 5 min,with a significant difference(IR at sAMY elevation/immediately after intervention=958±1391/476±1031,P<0.001).The bispectral indices,systolic blood pressure and pulse rates,which were measured at the same time,remained stable throughout the ESD.Forceful endoscopic insertion or over insufflation was performed during 22 of the 26 episodes.Release of the gastric wall tension and/or the administration of analgesic medication resulted in the immediate recovery of the elevated sAMY level,independent of body movement.CONCLUSION:By detecting twice the actual sAMY based on the preoperative level,the release of the gastric wall tension or the administration of analgesic agents should be considered.

Studies on Amylase and Degradation of Starch and Pigment of Tobacco Leaf During Process of Flue-Curing

The changes in the activity of amylase and amylase-isoenzyme and the degradation of starch and pigment of tobacco leaf during flue-curing were studied by using the electric- heated flue-curing barn designed and made by the Henan Agricultural University. The temperature and humidity of the barn were controlled automatically. The results indicated that starch in tobacco leaf decreased rapidly and leveled off after 48 h of curring, in the meantime, the content of soluble sugar increased accordingly and reached a peak at the stage of color-fixing. Both of them had a rapid-changing stage in the first 36 hours of yellowing. The changes of starch and soluble sugar contents had highly significant negative-correlation at 1 % level (rNC89 = -0.8962**, rYY85 = -0.9704**). The activity of amylase increased with the proceeding of curing and reached a peak after 36 hours of curing, then decreased. But the activity of amylase kept at a high level when the humidity of curing-environment was very low, even if the tobacco leaf had been dried. The rapid degradation of starch showed a significantly negative correlation with the increase of activity of amylase at 5 % level (rNC89 = -0.8495*, rYY85 = -0.7839*). The degradation of starch and pigment had the same regulation and had highly significant correlation at 1 % level (rNC89= 0.9649**, rYY85= 0.9428**). There were mainly three amylase-isoenzyme bands -A, B, C respectively, in tobacco leaf during flue curing. They were identified as α-AMY, β-AMY, R-AMY, and the activity of β-AMY was the highest. The changes in amylase activity and contents of starch and pigment were affected by the tobacco leaf moisture and environmental humidity during curing.

Effects of Cadmium Stress on Seed Germination, Seedling Growth and Seed Amylase Activities in Rice (Oryza sativa)

Two rice varieties, Xiushui 110 with high cadmium (Cd) tolerance and Xiushui 11 with low Cd tolerance were used to study the effects of Cd stress on seed germination, seedling growth and amylase activities. The low cadmium concentration had little effect on seed germination rate. However, cadmium stress could significantly inhibit plumule and radicle growth, especially for radicle growth. Germination index, vigour index, radicle length and amylase activities of Xiushui 11 decreased more significantly with the increasing cadmium level compared with Xiushui 110. The cadmium content in seedlings of Xiushui 11 was higher than that in Xiushui 110 when the cadmium concentration exceeded 5 μmol/L, which caused lower mitotic index in root tips and amylase activities, and more serious cadmium toxicity in Xiushui 11.

Low serum amylase and obesity, diabetes and metabolic syndrome: A novel interpretation

For the last decade, low serum amylase(hypoamylasemia) has been reported in certain common cardiometabolic conditions such as obesity, diabetes(regardless of type), and metabolic syndrome, all of which appear to have a common etiology of insufficient insulin action due to insulin resistance and/or diminished insulin secretion. Some clinical studies have shown that salivary amylase may be preferentially decreased in obese individuals, whereas others have revealed that pancreatic amylase may be preferentially decreased in diabetic subjects with insulin dependence. Despite this accumulated evidence, the clinical relevance of serum, salivary, and pancreatic amylase and the underlying mechanisms have not been fully elucidated. In recent years, copy number variations(CNVs) in the salivary amylase gene(AMY1), which range more broadly than the pancreatic amylase gene(AMY2A and AMY2B), have been shown to be well correlated with salivary and serum amylase levels. In addition, low CNV of AMY1, indicating low salivary amylase, was associated with insulin resistance, obesity, low taste perception/satiety, and postprandial hyperglycemia through impaired insulin secretion at early cephalic phase. In most populations, insulin-dependent diabetes is less prevalent(minor contribution) compared with insulin-independent diabetes, and obesity is highly prevalent compared with low body weight. Therefore, obesity as a condition that elicits cardiometabolic diseases relating to insulin resistance(major contribution) may be a common determinant for low serum amylase in a general population. In this review, the novel interpretation of low serum, salivary, and pancreas amylase is discussed in terms of major contributions of obesity, diabetes, and metabolic syndrome.

Characterization of a Novel Mesophilic Bacterial Amylase Secreted by ZW2531-1, a Strain Newly Isolated from Soil

A novel mesophilic bacterial amylase, named oligosaccharide-producing multifunctional amylase（OPMA）, was discovered and characterized. OPMA is an extracellular enzyme secreted by ZW253 1-1, a strain newly isolated from Chinese soil. It could be purified to homogeneity from the culture supernatant of ZW2531-1 by 30%-60% saturated ammonium sulfate precipitation, followed by twice Sephadex gel filtration chromatography. OPMA is a 66 kDa protein based on SDS-PAGE and has an isoelectric point（p/） at pH=5.3 by Isoelectric focusing electrophoresis（WE）. It only catalyzes the degradation of starch, rather than other alpha-l,4- and/or 1,6-glucan polysaccbarides such as fl-cyclomaltodextrin and pullulan. OPMA degraded starch to produce several oligosccharides including maltose, maltotriose, and isomaltotriose as the major end-products, and perhaps other oligosaccharides such as isomalto- tetraose, rather than glucose. OPMA exhibited optimal catalytic activity at a reaction temperature of 50 ℃ and pH=6.0, as determined by orthogonal test. Under the optimal reaction conditions, purified OPMA bad a specific activity of 13.75 U/rag. These findings suggest that OPMA could be used for the production of some oligosaccharides beneficial to the food industry and medicine.

Chylous ascites secondary to hyperlipidemic pancreatitis with normal serum amylase and lipase

A 54-year old man with a family history of hyperlipidemia was admitted with a 12 h history of severe generalized abdominal pain associated with nausea, vomiting and abdominal distension. Examination of the abdomen revealed tenderness in the periumblical area with shifting dullness. Serum pancreatic amylase was 29 IU/L and lipase 44 IU/L, triglyceride 36.28 mmol/L. Ultrasound showed ascites. CT of the abdomen with contrast showed inflammatory changes surrounding the pancreas consistent with acute pancreatitis. Ultrasound (US) guided abdomen paracentesis yielded a milky fluid with high triglyceride content consistent with chylous ascites. The patient was kept fasting and intravenous fluid hydration was provided. Meperidine was administered for pain relief. On the following days the patient’s condition improved and he was gradually restarted on a low-fat diet, and fat lowering agent (gemfibrozil) was begun, 600 mg twice a day. On d 14, abdomen US was repeated and showed fluid free peritoneal cavity. The patient was discharged after 18 d of hospitalization with 600 mg gemfibrozil twice a day. At the time of discharge, the fasting triglyceride was 4.2 mmol/L. After four weeks the patient was seen in the clinic, he was well.

Supplementation of amylase combined with glucoamylase or protease changes intestinal microbiota diversity and benefits for broilers fed a diet of newly harvested corn

Background: The effect of amylases combined with exogenous carbohydrase and protease in a newly harvested corn diet on starch digestibility, intestine health and cecal microbiota was investigated in broiler chickens.Methods: Two hunderd and eighty-eight 5-day-old female chickens were randomly divided into six treatments: a newly harvested corn-soybean meal diet(control); control supplemented with 1,500 U/g α-amylase(Enzyme A);Enzyme A + 300 U/g amylopectase + 20,000 U/g glucoamylase(Enzyme B); Enzyme B + protease 10,000 U/g(Enzyme C); Enzyme C + xylanase 15,000 U/g(Enzyme D); and Enzyme D + cellulase 200 U/g + pectinase 1,000 U/g(Enzyme E). Growth performance, starch digestibility, digestive organ morphology, and intestinal microbiota were evaluated in the birds at 16 and 23 d of age.Results: Compared with the control diet, supplementation with Enzyme A significantly decreased ileum lesion scoring at 16 d of age(P < 0.05); supplementation with Enzyme B or Enzyme C showed positive effects on ileal amylopectin and total starch digestibility(P < 0.05); Broilers fed with a diet supplemented with Enzyme D had a tendency to decrease body weight gain at 23 d. Enzyme E supplementation improved lesion scoring of jejunum and ileum at 16 d(P < 0.05), and increased ileal amylopectin or total starch digestibility at 23 d(P < 0.05).Supplementation of enzymes changed cecal microbiota diversity. High numbers of Campylobacter, Helicobacter and Butyricicoccus, Anaerostipes and Bifidobacterium, Sutterella and Odoribacter were the main genera detected in supplementations with Enzymes B, C, D, and E respectively.Conclusions: Supplementation with amylase combined with glucoamylase or protease showed a beneficial effect on starch digestibility and intestinal microbiota diversity, and increased growth of broilers fed with newly harvested corn.

Combination of two-hour post-endoscopic retrograde cholangiopancreatography amylase levels and cannulation times is useful for predicting post-endoscopic retrograde cholangiopancreatography pancreatitis

AIM To estimate the efficacy of 2 h post-endoscopic retrograde cholangiopancreatography(ERCP) serum amylase levels and other factors for predicting postERCP pancreatitis.METHODS This was a retrospective,single-center cohort study of consecutive patients who underwent ERCP from January 2010 to December 2013.Serum amylase levels were measured 2 h post-procedure,and patient- and procedure-related pancreatitis(PEP) risk factors wereanalyzed using a logistic model.RESULTS A total of 1520 cases(average age 72 ± 12 years,60% male) were initially enrolled in this study,and 1403 cases(725 patients) were ultimately analyzed after the exclusion of 117 cases.Fifty-five of these cases developed PEP.We established a 2 h serum amylase cutoff level of two times the upper limit of normal for predicting PEP.Multivariate analysis revealed that a cannulation time of more than 13 min [odds ratio(OR) 2.28,95%CI:1.132-4.651,P=0.0210] and 2 h amylase levels greater than the cutoff level(OR=24.1,95%CI:11.56-57.13,P<0.0001) were significant predictive factors for PEP.Forty-seven of the 55 patients who developed PEP exhibited 2 h amylase levels greater than the cutoff level(85%),and six of the remaining eight patients who developed PEP(75%) required longer cannulation times.Only 2 of the 1403 patients(0.14%) who developed PEP did not exhibit concerning 2 h amylase levels or require longer cannulation times.CONCLUSION These findings indicate that the combination of 2 h post-ERCP serum amylase levels and cannulation times represents a valuable marker for identifying patients at high risk for PEP.

Diagnostic value of drain amylase for detecting intrathoracic leakage after esophagectomy

AIM:To investigate the value of elevated drain amylase concentrations for detecting anastomotic leakage(AL) after minimally invasive Ivor-Lewis esophagectomy(MIILE).METHODS:This was a retrospective analysis of prospectively collected data in two hospitals in the Netherlands. Consecutive patients undergoing MI-ILE were included. A Jackson-Pratt drain next to the dorsal side of the anastomosis and bilateral chest drains were placed at the end of the thoracoscopic procedure. Amylase levels in drain fluid were determined in all patients during at least the first four postoperative days. Contrast computed tomography scans and/or endoscopic imaging were performed in cases of a clinically suspected AL. Anastomotic leakage was defined as any sign of leakage of the esophago-gastric anastomosis on endoscopy,re-operation,radiographic investigations,post mortal examination or when gastro-intestinal contents were found in drain fluid. Receiver operator characteristic curves were used to determine the cut-off values. Sensitivity,specificity,positive predictive value,negative predictive value,risk ratio and overall test accuracy were calculated for elevated drain amylase concentrations.RESULTS:A t o t a l o f 8 9 p a t ie n t s w e re in c lu d e d between March 2013 and August 2014. No differences in group characteristics were observed between patients with and without AL,except for age. Patients with AL were older than were patients without AL(P = 0.01). One patient(1.1%) without AL died within 30 d after surgery due to pneumonia and acute respiratory distress syndrome. Anastomotic leakage that required any intervention occurred in 15 patients(16.9%). Patients with proven anastomotic leakage had higher drain amylase levels than patients without anastomotic leakage [median 384 IU/L(IQR 34-6263) vs median 37 IU/L(IQR 26-66),P = 0.003]. Optimal cut-off values on postoperative days 1,2,and 3 were 350 IU/L,200 IU/L and 160 IU/L,respectively. An elevated amylase level was found in 9 of the 15 patients with AL. Five of these 9 patients had early elevations of their amylase levels,with a median of 2 d(IQR 2-5) before signs and symptoms occurred.CONCLUSION:Measurement of drain amylase levels is an inexpensive and easy tool that may be used to screen for anastomotic leakage soon after MI-ILE. However,clinical validation of this marker is necessary.

Drainage fluid and serum amylase levels accurately predict development of postoperative pancreatic fistula

AIM To investigate potential biomarkers for predicting postoperative pancreatic fistula( POPF) after pancreaticoduodenectomy(PD).METHODS We prospectively recruited 83 patients to this study. All patients underwent PD(Child's procedure) at the Division of Hepatobiliary and Pancreas Surgery at the First Bethune Hospital of Jilin University between June 2011 and April 2015. Data pertaining to demographic variables, clinical characteristics, texture of pancreas, surgical approach, histopathological results, white blood cell count, amylase and choline levels in the serum, pancreatic/gastric drainage fluid, and choline and amylase levels in abdominal drainage fluid were included in the analysis. Potential correlations between these parameters and postoperative complications such as, POPF, acute pancreatitis, hemorrhage, delayed gastric emptying, and biliary fistula, were assessed. RESULTS Twenty-eight out of the 83(33.7%) patients developed POPF. The severity of POPF was classified as Grade A in 8(28%) patients, grade B in 16(58%), and grade C in4(14%), according to the pancreatic fistula criteria. On univariate and multivariate logistic regression analyses, higher amylase level in the abdominal drainage fluid on postoperative day(POD)1 and higher serum amylase levels on POD4 showed a significant correlation with POPF(P < 0.05). On receiver operating characteristic curve analysis, amylase cut-off level of 2365.5 U/L in the abdominal drainage fluid was associated with a 78.6% sensitivity and 80% specificity [area under the curve(AUC): 0.844; P = 0.009]. A cut-off serum amylase level of 44.2 U/L was associated with a 78.6% sensitivity and 70.9% specificity(AUC: 0.784; P = 0.05).CONCLUSION Amylase level in the abdominal drainage fluid on POD1 and serum amylase level on POD4 represent novel biomarkers associated with POPF development.

Heterologous Expression of Amylase Gene from Saccharomycopsis fibuligera in an Industrial Strain of Saccharomyces cerevisiae

An α-amylase encoding gene was amplified by polymerase chain reaction from Saccharomycopsis fibuligeru and inserted into a shuttle vector YEp352,together with the yeast phosphoglycerate kinase 1 prumoter and a-factor signal gene. The recombinant expression plasmid pLA8α was transformed into an industrial strain of Saccharomycopsis cerevisiae Sc-11. The activity of the α-amylase produced by the transformant Sc-11-pLA8α was 6.3 U/mL and the starch utilization rate in YPS medium was 42%. The purified amylase was analyzed by SDS-PAGE,showing a molecular weight of 55×10^3 protein band. Furthermore, the residual sugar, ethanol and some volatile compounds in the fermented worts under simulating brewing conditions were determined by chromatographic analyses. The fermentation characteristics of Sc-ll-pLA8α were similar to that of Sc-ll and only minor changes in the concentration of flavor compounds could be observed.

A marked enhancement in production of amylase by Bacillus amyloliquefaciens in flask fermentation using statistical methods

A total of 126 bacterial strains were isolated from soil samples. Among them, 11 isolates were found positive for amylase production. Strain YL produced the largest zone of clearance on plate assay. The isolate YL was identified as Bacillus sp. based on morphological and physiochemical characterization. According to 16S rRNA gene sequencing data, the closest phylogenetic neighbor of strain YL was Bacillus amyloliquefaciens （99.54%）. After that, an optimization of culture conditions was carried out for the improvement of a-amylase production. Response surface methodology （RSM） was applied to evaluate the effect of medium components including wheat bran, cottonseed extract, yeast extract, starch, NaC1 and CaCl2. Three variables （wheat bran, cottonseed extract, and starch）, which were identified to significantly affect amylase production by Plackett-Burman design were further optimized using response surface methodology of Box-Behnken design （BBD）. The optimal concentrations estimated for each variable related to the maximum of amylase activity （86 kU/mL） were 10.80 g/L wheat bran, 9.90 g/L cottonseed extract, 0.5 g/L starch, 2.0 g/L yeast extract, 5.00 g/L NaCl and 2.00 g/L CaC12. The fermentation using optimized culture medium allowed a significant increase in amylase production （by 3-fold）. The improvement in the a-amylase production after optimization process can be considered adequate for large-scale applications.