Extraction, Partial Characterization, and Storage Stability of β‐Glucosidase from Propolis
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Abstract: Extraction and assay conditions for β‐glucosidase from propolis were optimized. Highest enzyme activity was obtained in a citric acid‐disodium hydrogen phosphate buffer at pH 6.0 with 2.5% insoluble polyvinylpyrrolidone at incubation temperature of 57 °C. β‐Glucosidase activities were found in all freshly harvested propolis while β‐glucosidase activities were scarcely present in the randomly bought propolis. Propolis was stored at –20 °C and 4 °C for 3 mo with almost no loss of β‐glucosidase activity, but at room temperature the activity decreased exponentially with the increase of storage time. These results indicated that the activity of β‐glucosidase could be a candidate for propolis‐freshness index. β‐Glucosidase from propolis was capable of hydrolyzing p‐ nitrophenyl‐β‐D‐glucoside and p‐ nitrophenyl‐β‐D‐galactoside, but lacked activity toward p‐ nitrophenyl‐β‐D‐glucuronide, p‐ nitrophenyl‐β‐D‐cellobioside, amygdalin, cellobiose, and gentiobiose. These results were consistent with the hypothesis that flavonoid glucosides were hydrolyzed by β‐glucosidase during propolis collection and processing and provided a possible explanation for why some flavonoid biosides (that is, rutin and isorhamnetin‐3‐O‐rutinoside) exist in propolis. Practical Application: β‐Glucosidase activity was detected and partial characterization of the enzyme was determined in propolis. The enzyme activity decreased exponentially with the increase of storage time at room temperature, which suggested that the activity of β‐glucosidase could be regarded as a freshness index of propolis. The research will be useful for studying the chemical constituents of propolis.Keywords:
Propolis
Abstract α‐Amylase of the thermophilic actinomycete Thermomonospora vulgaris was partially purified. Maximal enzyme activity was obtained at 60 °C and pH 6.0. K M value was 1.4%. The effect of some metal salts on enzyme activity was studied. Enzyme activity was inhibited by KCN, EDTA, and iodoacetate. Inhibition by EDTA was completely nullified by CaCl 2 , but the inhibition by iodoacetate was not overcome by 2‐mercaptoethanol. Exposure of the enzyme to pH 7.0 and 9.0 for 2hr. did not affect the enzyme, but exposure to pH 3.0 for few minutes completely inactivated the enzyme. Exposure of the enzyme to 60 °C resulted in an appreciable inactivation and exposure to 80 °C completely inactivated the enzyme. Addition of CaCl 2 , 2‐mercaptoethanol, or enzyme substrate did not stabilize the 60 °C exposed enzyme. However, bovine serum albumin had a protective effect when the enzyme was exposed to 60 °C but not to 80 °C. The enzyme was stable in the presence of 8 M urea.
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Two ethanol and supercritical carbon dioxide( CO2) extraction methods were used to produce propolis extracts from 9 districts and 16 varieties propolis samples in this study. The yield and total flavonoid content of propolis extracts were determined correspondingly. In addition,the propolis area differentiation was evaluated by data statistical analysis. By comparing two extraction methods,the results showed that the total flavonoid content of propolis extracts produced by the supercritical CO2extraction method was significantly lower than that produced by the ethanol extraction method. The propolis area differentiation results showed that the propolis grown in Hebei and Yunnan regions had generally higher yields and total flavonoid contents,the propolis developed at Hubei,Shandong,Hunan and Hainan areas had moderate yields and total flavonoid contents,and the propolis grown in Heilongjiang region had generally lower yield and total flavonoid content. Moreover,the total flavonoid contents of propolis extracts from different varieties grown in Henan and Sichuan regions showed significant differences. Therefore,this research suggested that the propolis production enterprises fixed the raw materials collecting area,variety and extraction method in order to guarantee the propolis product quality stability.
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In this study, we evaluated optimum condition of enzyme with pH and temperature for preparation of microfibillated cellulose(MFC). Well-known endo-glucanase, three enzymes were used and CMC was used for substrate. Enzyme activity was evaluated using DNS method and absorbance with UV/VIS spectrophotometer. The enzyme shown the greatest activity was reacted with pulps at optimum condition for 1 hour and treated pulps beated until 100 mL CSF. Enzyme B and Enzyme L was the higher enzyme activity below 0.1% concentration and Enzyme N was the lowest enzyme activity. At various pH and temperature conditions, enzyme activity of Enzyme B was higher than the others at the same concentration. Especially enzyme activity at 50℃ of Enzyme B was almost not changed over pH 6.0. Optimum condition of three enzyme was pH 6 or pH 7 and 50℃or 60℃. Also beating efficiency of enzyme treated pulps with Enzyme B is 55.6%.
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e studied the effect of temperature, enzyme concentration and pH on enzyme activity. The enzyme we studied was hydrogen peroxidase from a cow. The reaction converted hydrogen peroxide to water and oxygen and oxygen production was used as a measure of enzyme activity. We studied enzyme activity at temperatures of 9 C, 37 C, 41 C. It showed that at 9 C, there was almost no activity. The activity at 41 was 1.5X the activity at 37. The enzyme was tested using 1⁄2X, 1X and 2X enzyme concentration. Significant enzyme activity was seen at 1x enzyme concentration and the 1⁄2X enzyme concentration trial showed almost the same activity. The activity at 2X enzyme concentration was approximately 2 times that at 1X enzyme concentration. Finally, we tested the effect of pH on the enzyme with pH 7, pH 1 and pH 11. The enzyme activity was highest at neutral pH (7) and showed only 1/3 enzyme activity at pH 11while pH of 1 showed no enzyme activity. Introduction: Enzymes are catalysts or chemical agents that speed up chemical reactions without being consumed by the reaction (Reece et al. 2010). Most enzymes are proteins that function to reduce activation energy in chemical reactions (Petersen and Anderson 2005). Enzymes work on reactants called substrate; the enzyme attaches to the substrate and then the enzyme converts the substrate to products while the enzyme remains unaffected (Reece et al. 2010). Enzyme activity can be affected by environmental factors (Petersen and Anderson 2005). Temperature is one environmental factor that can affect enzyme activity (Conant 2012). Another factor that affects enzymes is pH (Leake and Read 1990). In this lab, we will be studying the effects of temperature, enzyme concentration and pH on the enzyme, hydrogen peroxidase that is found in all aerobic cells and functions to decompose hydrogen peroxide (Petersen and Anderson 2005). Materials and methods: A strip of filter paper was dipped into a cow liver homogenate for 10 seconds and then placed in a chamber. 20 mL of 1.5% H2O2 was added to the chamber and the chamber was plugged with a stopper and then placed in a water bath. After 5 minutes, a 50 mL graduated cylinder filled with water was overturned over the chamber to flow the H2O2. Measurements of oxygen released were taken at 5 second intervals for one minute for each trial. This experiment was done with three different variables: Temperature, enzyme concentration and pH. The temperatures tested were: 9 C, 37 C, 41 C. The enzyme concentration was done using 1⁄2 strip, 1 strip and 2 strips of filter paper (representing 1⁄2X, 1X and 2X enzyme concentration) keeping the temperature at 37 C. The pH test was done using 1 strip with 5 drops of 50% HCl added to the H2O2 to produce a pH of 1. Another pH test was performed by adding 5 drops of 50% NaOH to the H2O2 to yield a pH of 11. The third trial was kept at pH 7. The temperature for the pH trials was at 37 C. W 1 Eed: Factors Affecting Enzyme Activity Produced by The Berkeley Electronic Press, 2013
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alpha-Amylase of the thermophilic actinomycete Thermomonospora vulgaris was partially purified. Maximal enzyme activity was obtained at 60degreeC and pH 6.0. KM value was l.4%. The effect of some metal salts on enzyme activity was studied. Enzyme activity was inhibited by by KCN, EDTA, and iodoacetate. Inhibition by EDTA was completely nullified by CaCl2, but the inhibition by iodoacetate was not overcome by 2-mercaptoethanol. Exposure of the enzyme to pH 7.0 and 9.0 for 2 hr. did not affect the enzyme, but exposure to pH 3.0 for few minutes completely inactivated the enzyme. Exposure of the enzyme to 60degreeC resulted in an appreciable inactivation and exposure to 80degreeC completely inactivated the enzyme. Addition of CaCl2, 2-mercaptoethanol, or enzyme substrate the 60degreeC exposed enzyme. However, bovine serym albumin had a protective effect when the enzyme was exposed to 60degreeC but not to 80degreeC. The enzyme was stable in the presence of 8 M urea.
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A strain producing fibrinolytic enzyme was isolated from starter.The strain was identified as Rhizopus micro-sporus var.tuberosus based on the morphological and ITS sequence.The enzymatic properties of the fibrinolytic enzyme secreted by the strain were determined.The properties tests suggest that the optimum temperature and pH were 37,℃ and 7.0r,espectively.The enzyme retained 95% of its initial activity by retaining for 4 h at 37,℃,and when it was treated at 57 ℃ for 4 h,the enzyme was still remained of activity.The enzyme had a wide range of pH adaptabilityi,t still remained of part of activity when pH was adjusted to a level(pH3.4);and it remained enzyme activity in between the range of pH 6.0-8.0.Zn2+ and Cu2+ have a very clear function to restrain the enzyme activity whereas Na+,Ca2+,Mn2+ showed obvious stimu-lation on the enzyme.The results show that the enzyme is stable in the artificial intestinal juicei,t retained 80% of its initial activity by retaining for 4 h at 37,℃.It may be developed to injection used in clinical research.
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Abstract Propolis, a complex substance of resinous, is produced by honeybees. Propolis contains a number of flavonoid compounds endowed with antimicrobial and antioxidant capabilities. The main chemical constituents and physicochemical properties of propolis are depended on many factors including geographic origin. This study was undertaken to evaluate antibacterial, antibiofilm, and antioxidant activities of propolis collected from Kalimantan, Indonesia. The phytochemical properties of propolis were identified by total phenolic contents, total flavonoid contents, and FTIR analysis. Propolis was extracted using 96% ethanol, 70% ethanol, and n-hexane solvents. Moreover, the bioactivity capacity of propolis extracts were observed through antioxidant and antibacterial activities. The results showed that propolis extracts exhibited varying degrees of phenolic content, flavonoid contents, antibacterial, and antiradical scavenging properties. The highest value of total flavonoid content was obtained on 96% ethanol propolis extract as well as its remarkable antibacterial and antiradical scavenging activities.
Propolis
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