Khảo sát điều kiện trích ly và tinh sạch lectin từ đậu ma (Pueraria phaseoloides) bằng sắc ký ái lực

2021 ◽  
Vol 57 (6) ◽  
pp. 151-159
Author(s):  
Văn Song Toàn Võ ◽  
Khoa Nguyên Trần ◽  
Thị Thu Xương Kim ◽  
Thị Bảo Trân Nguyễn ◽  
Hoàng Thịnh Nhan ◽  
...  
Keyword(s):  
Ammonium Sulfate ◽  
Pueraria Phaseoloides ◽  
Hemagglutination Assay ◽  
Sds Page

Nghiên cứu được thực hiện nhằm xác định điều kiện trích ly và tinh sạch lectin từ đậu ma Pueraria phaseoloides. Lectin đậu ma được trích ly cùng với dung dịch NaCl 0,9% ở các tỷ lệ (w/v), thời gian và nhiệt độ ủ khác nhau. Dịch chiết thô được tinh sạch bằng phương pháp tủa phân đoạn với muối ammonium sulfate, tiếp theo là sắc ký ái lực trên gel Sepharose D-galactose để cải thiện độ tinh sạch. Kết quả cho thấy lectin đậu ma đạt hiệu quả trích ly tối ưu với hoạt tính đặc hiệu đạt là 1.579 HAA (Hemagglutination assay)/mg ở tỉ lệ với dung môi trích ly là 1:4 (w/v), tại 50oC, trong 10 phút. Dịch trích lectin đậu ma sau khi tủa phân đoạn ở nồng độ muối 40% - 50% cho hiệu suất thu hồi 35,4% với độ tinh sạch tăng 6,38 lần so với dịch trích thô; trong khi phân đoạn F1 từ sắc ký ái lực cho hiệu suất thu hồi 9,85% với độ tinh sạch tăng 16,2 lần. Kết quả điện di SDS-PAGE xuất hiện hai băng protein có khối lượng phân tử 66,0 kDa và 56,0 kDa.

scholarly journals Purification and Characterization ofα -l-Arabinopyranosidase andα -l-Arabinofuranosidase from Bifidobacterium breve K-110, a Human Intestinal Anaerobic Bacterium Metabolizing Ginsenoside Rb2 and Rc

2003 ◽  
Vol 69 (12) ◽  
pp. 7116-7123 ◽  
Author(s):  
Ho-Young Shin ◽  
Sun-Young Park ◽  
Jong Hwan Sung ◽  
Dong-Hyun Kim
Keyword(s):  
Ammonium Sulfate ◽  
Column Chromatography ◽  
Specific Activity ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Bifidobacterium Breve ◽  
Ammonium Sulfate Fractionation ◽  
Apparent Homogeneity ◽  
Sds Page ◽  
Ginsenoside Rb2

ABSTRACT Two arabinosidases, α-l-arabinopyranosidase (no EC number) and α-l-arabinofuranosidase (EC 3.2.1.55), were purified from ginsenoside-metabolizing Bifidobacterium breve K-110, which was isolated from human intestinal microflora. α-l-Arabinopyranosidase was purified to apparent homogeneity, using a combination of ammonium sulfate fractionation, DEAE-cellulose, butyl Toyopearl, hydroxyapatite Ultrogel, QAE-cellulose, and Sephacryl S-300 HR column chromatography, with a final specific activity of 8.81 μmol/min/mg.α -l-Arabinofuranosidase was purified to apparent homogeneity, using a combination of ammonium sulfate fractionation, DEAE-cellulose, butyl Toyopearl, hydroxyapatite Ultrogel, Q-Sepharose, and Sephacryl S-300 column chromatography, with a final specific activity of 6.46 μmol/min/mg. The molecular mass ofα -l-arabinopyranosidase was found to be 310 kDa by gel filtration, consisting of four identical subunits (77 kDa each, measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis [SDS-PAGE]), and that ofα -l-arabinofuranosidase was found to be 60 kDa by gel filtration and SDS-PAGE. α-l-Arabinopyranosidase and α-l-arabinofuranosidase showed optimal activity at pH 5.5 to 6.0 and 40°C and pH 4.5 and 45°C, respectively. Both purified enzymes were potently inhibited by Cu2+ and p-chlormercuryphenylsulfonic acid.α -l-Arabinopyranosidase acted to the greatest extent on p-nitrophenyl-α-l-arabinopyranoside, followed by ginsenoside Rb2. α-l-Arabinofuranosidase acted to the greatest extent on p-nitrophenyl-α-l-arabinofuranoside, followed by ginsenoside Rc. Neither enzyme acted on p-nitrophenyl-β-galactopyranoside or p-nitrophenyl-β-d-fucopyranoside. These findings suggest that the biochemical properties and substrate specificities of these purified enzymes are different from those of previously purified α-l-arabinosidases. This is the first reported purification ofα -l-arabinopyranosidase from an anaerobic Bifidobacterium sp.

scholarly journals PRODUCTIVITY Pueraria phaseoloides var. Javanica FERTILIZED WITH MANURE FROM CATTLE OFFERED CONCENTRATE CONTAINNING DIET SUPPLEMENTED WITH AMMONIUM SULFAT

Pastura ◽  
2019 ◽  
Vol 8 (1) ◽  
pp. 39
Author(s):  
Tjok. Istri Putri ◽  
Ni Putu Mariani
Keyword(s):  
Ammonium Sulfate ◽  
Weight Ratio ◽  
Dry Weight ◽  
Optimal Dosage ◽  
Pueraria Phaseoloides ◽  
Leaf Color ◽  
Randomized Design ◽  
Number Of Leaves ◽  
The Difference ◽  
Number Of Branches

This study aims to determine the optimal dosage of manure derived from cattle fed concentrated rations supplemented ammonium sulfate which can increase the productivity of Pueraria phaseoloidesvar .Javanica. The design used in this experiment was Completely Randomized Design (RAL) with 4 treatments and 5 replications. The four treatments were fertilized with doses of 10, 15, 20 and 25 tons / ha, respectively for the treatment of A, B, C and D, so overall there were 20 pot experiments. The study was conducted for 12 weeks. Observed variables were plant length, number of leaves, number of branches, dry weight (DW) of leaves, DW of stem DW, total BK, leaf color, leaf stem ratio (LSR) and leaf weight ratio”(LWR). The results showed that legumes treated A B, C and D produced high crop, leaf area, leaf color, LSR and LWR were not significantly different (P> 0.05). The treated legume C. resulted in a significantly higher leaf number than the treated legume A, B and D (P <0.05). The number of branches and DW leaves on legumes treated with C were significantly higher (P <0.05) than those receiving treatment A. and were not significantly different with legumes treated with B and D (P> 0.05). The DW of stem and total DW production from legume treated C were significantly higher with those treated with A and B (P <0.05) and the difference was not significant with those treated with D (P> 0.05). From the results of the study it can be concluded that legume treated with C or fertilized with manure derived from cattle fed concentrate containning diet supplemented ammonium sulfate at a dose of 20 tons / ha yield of leguminous productivity Peuraria phaseoloides var. javanica maximum. Keywords: manure, pueraria phaseoloides, productivity.

scholarly journals ISOLATION OF PROTEIN FROM THE SPINE VENOM OF PTEROIS VOLITANS FOUND IN THE INDONESIAN OCEAN, USING A HEATING PROCESS, FOR ANTICANCER, ANTIRETROVIRAL, ANTIBACTERIAL, AND ANTIOXIDANT ASSAYS

2021 ◽  
pp. 53-58
Author(s):  
FERA IBRAHIM ◽  
MUHAMAD SAHLAN ◽  
MIKAEL JANUARDI GINTING ◽  
DIAH KARTIKA PRATAMI ◽  
HERI HERMANSYAH ◽  
...  
Keyword(s):  
Anticancer Activity ◽  
Ammonium Sulfate ◽  
Heating Temperature ◽  
Heating Process ◽  
Pterois Volitans ◽  
Isolation And Purification ◽  
E Coli ◽  
Sds Page ◽  
Agar Diffusion Test ◽  
Dpph Method

Objective: This research investigates the antibacterial, anticancer, antioxidant, and antiretroviral activities of the lionfish spine poison extract. Methods: Isolation and purification of the phospholipase A2 (PLA2) protein obtained from the spine poison were conducted through the following stages, including, extraction of the venom by sonication, heating, and purification using gradual saturation levels of ammonium sulfate. Furthermore, the purity and concentration of PLA2 were analyzed using the Lowry test and Marinetti’s method, respectively, while its protein content was ascertained through SDS-PAGE. Toxicity was then evaluated employing the brine shrimp lethality test (BSLT), and its anticancer activity was assessed in human cervical carcinoma cells (HeLa cells). Finally, its antioxidant, antibacterial, and antiretroviral activities were analyzed using the DPPH method, agar diffusion test against Salmonella sp. and E. coli, and SRV-2 and RT-qPCR tests, respectively. Results: The protein demonstrated 37.79% inhibition for anticancer activity, IC50 1312 ppm for antioxidant activity, 98.81%, and 89.28% inhibition of E. coli and Salmonella sp. respectively for antibacterial activity and 98.13% inhibition for antiretroviral activity. Conclusion: It can be concluded that lionfish (Pterois volitans) has the potential to be developed as an antioxidant, anticancer, antibacterial, and antiretroviral agent. Furthermore, the pharmacological activity of its spine venom was determined by isolating PLA2 protein from its extract, using an optimum heating temperature of 70 °C and an ammonium sulfate saturation level of 80%.

scholarly journals Characterization of Gelatin Profile of Chicken Broiler (Gallus domestica) Bone Using SDS-PAGE Electrophoresis

ALCHEMY ◽  
2019 ◽  
Vol 7 (1) ◽  
pp. 7 ◽  
Author(s):  
Dewi Yuliani ◽  
Dhienda Risa Awalsasi ◽  
Akyunul Jannah
Keyword(s):  
Ammonium Sulfate ◽  
Protein Concentration ◽  
Peptide Fragments ◽  
Strong Base ◽  
Chicken Bone ◽  
Sds Page ◽  
Hydrolysis Of ◽  
Β Chain ◽  
Page Electrophoresis

<p>Gelatin, a proteinaceous additive, is obtained from hydrolysis of collagen in the bone, hide and skin of animals. As natural product, gelatin has been applied in many industries with various functions. This study attempt to characterize gelatin profile of broiler chicken (<em>Gallus domestica</em>) using SDS-PAGE electrophoresis. The chicken bone was pretreated using a strong base, sodium hydroxide, producing type B gelatin. The gelatin was purified through precipitation using the variation of ammonium sulfate concentrations (40-70%) and dialysis using cellophane membrane. The purified gelatin was characterized through SDS-PAGE electrophoresis. Based on electrophoresis visualization, reduction of band intensity by ammonium sulfate 40% showed removal of small peptide fragments. The remained gelatin showed two major bands, α-chains and a β-chain with the respective molecular weight of ~135 and ~245 kDa. The protein content of the unpurified gelatin (E1) was 71.65±0.60 mg/L.  The purified E1 gelatins by 40-70% of ammonium sulfate addition contained 61.42±3.90, 60.45±1.36, 59.89±0.24, and 55.32±1.05 mg/L of protein concentration, respectively.</p><p> </p><p>Keywords: chicken bone, gelatin profile, protein electrophoresis</p>

Preliminary Purification and Distribution of Lipoxygenase in Grass Carp

2012 ◽  
Vol 550-553 ◽  
pp. 1468-1472 ◽  
Author(s):  
Jian Hui Wang ◽  
Dong Min Liu ◽  
An Ni Zheng ◽  
Yong Le Liu ◽  
Qi Chen ◽  
...  
Keyword(s):  
Protein Content ◽  
Ammonium Sulfate ◽  
Grass Carp ◽  
Extraction Methods ◽  
Salting Out ◽  
Percentage Saturation ◽  
Preliminary Purification ◽  
Sds Page ◽  
Fish Product ◽  
And Storage

The lipoxygenase (LOX) in grass carp can catalyze the oxidation of polyunsaturated fatty acids even their esters, resulting in rancidity deterioration of grass carp and its products during the process and storage. Preliminary purification, quantitative determination and distribution of grass carp muscular LOX were investigated in the present study. LOX activity of the grass carp muscle was measured to be 35.17 KU/g and its molecular weight was 95.71 kDa identified by SDS-PAGE. Using previously reported extraction methods such as twice salting-out and once dialysis, the LOX protein content in the crude muscular LOX extraction is about 28%. When salted with graded saturation of ammonium sulfate, we found once salting-out was enough for LOX extraction and the effective percentage saturation of ammonium sulfate for LOX extraction was 40%. Under this condition, the LOX protein content in the crude muscular LOX extract was doubled. The LOX distribution in fish was gradually increased from skin and muscle, gill, viscera to blood, which indicates that it is very important to remove the viscera and bleed fish completely during fish product processing to decrease the risk of lipid oxidation caused by LOX.

Effects of Precipitation Methods on the Properties of Protease Extracted from Starfruit (Averrhoa carambola L.) of Different Maturity Index

2015 ◽  
Vol 12 (1) ◽  
pp. 11
Author(s):  
Normah Ismail ◽  
Ezzana Zuraini Zainuddin
Keyword(s):  
Ammonium Sulfate ◽  
Proteolytic Activity ◽  
Protein Concentration ◽  
Relative Activity ◽  
Raw Material ◽  
Maturity Index ◽  
Protease Production ◽  
Ammonium Sulphate Precipitation ◽  
Sds Page ◽  
Index 2

Proteases were extracted from starfruit at maturity Index 2 (unripe, light green) and Index 7 (very ripe, orange) and partially purified using acetone and 40% ammonium sulfate precipitations. Higher yield and proteolytic activity were observed for proteases purified using acetone than 40% ammonium sulfate. As for maturity index, yield and protein concentration of proteases from Index 2 were higher than those from Index 7. SDS-PAGE result showed intense bands for acetone proteases while a distinct band at 50 kDa was observed in all the proteases. Enzyme activity decreased during the seven days storage at 4°C with minimum relative activity of 70% achieved for acetone proteases at day seven. This study suggested that acetone precipitation is more effective method for purifying starfruit protease based on the yield and proteolytic activity compared to using 40% ammonium sulphate precipitation. In order to obtain higher protein concentration and proteolytic activity, starfruit at the unripe stage, Index 2 is a better raw material than Index 7 to be used for protease production. 

Preliminary Fractionation and Derivatives Changes of Myoglobin from Tilapia (O. niloticus♀×O. aureus♂) Dark Muscle with Ammonium Sulfate

2011 ◽  
Vol 393-395 ◽  
pp. 890-893
Author(s):  
Jian Wei Cen ◽  
Shu Xian Hao ◽  
Lai Hao Li ◽  
Xian Qing Yang ◽  
Hui Huang ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Ammonium Sulfate ◽  
Absorption Spectra ◽  
Dark Muscle ◽  
Ammonium Sulfate Fractionation ◽  
Sulfate Fractionation ◽  
Sds Page

Ammonium sulfate fractionation was implicated as one of essential steps to purified Myoglobin (Mb).We adopted SDS-PAGE analysis and absorption spectra scanning to demonstrate the effect of ammonium sulfate on Mb and its derivatives. The results shown that protein with the molecular weight above 37.8kDa were dominated in myoglobin extract of tilapia, which can be precipitated with ammonium sulfate. Mb molecular weight of tilapia is about 15.8 kDa, which can be collected by the treatment of ammonium sulfate with saturation between 60%-70% to remove unwanted protein. The absorption spectra show that Mb derivatives could be transferred to met-myoglobin after preliminary fractionating with ammonium sulfate.

scholarly journals Bacteriostatic activity of con a lectin from Canavalia ensiformis

2013 ◽  
Vol 1 (04) ◽  
pp. 59-63 ◽  
Author(s):  
S. R. Kulkarni ◽  
V. J. Tayade
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antibacterial Activity ◽  
Bacteriostatic Activity ◽  
Hemagglutination Activity ◽  
Con A ◽  
Plate Method ◽  
Hemagglutination Assay ◽  
Canavalia Ensiformis ◽  
Sds Page

The aim of this work was to explore the therapeutic applications of Con A lectin from Canavalia ensiformis and to explore its antibacterial activity. Activity of lectin was quantified by their ability to agglutinate erythrocytes using Hemagglutination assay. Characterization and purity of Con A lectin was evaluated by using SDS-PAGE analysis. The reversal of hemagglutination activity of lectin was evaluated by using the sugars namely; mannose, galactose, lactose, fructose, glucose. The antibacterial activity of lectins was tested against Streptococcus mutans, Staphylococcus aureus, Bacillus subtilis, Escherichia coli using pour plate method. Amoxycillin was used as standard. At 250mg/ml concentration Con A lectin showed good bacteriostatic activity.

The Purification of Grifolan Synthase and the Determination of its Molecular Weight

2012 ◽  
Vol 550-553 ◽  
pp. 1222-1228
Author(s):  
Peng Li ◽  
Wei Guo Wang ◽  
Yu Jia Cui ◽  
Yong Liang Zhao ◽  
Ya Nan Gu ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Ammonium Sulfate ◽  
Crude Extract ◽  
Polyacrylamide Gel Electrophoresis ◽  
Grifola Frondosa ◽  
Buffer Solution ◽  
Native Page ◽  
Purification Method ◽  
Sds Page ◽  
And Function

Purpose To search for a purification method of Grifolan Synthase crude extract from the mycelium of Grifola frondosa and determine its molecular weight. Methods Grifolan Synthase crude extract from the mycelium of Grifola frondosa was purified by recrystallization and native-PAGE. The process is as follows: Grifolan Synthase crude extract was dissolved in buffer solution (pH7.0). 10ml Grifolan Synthase crude extract solution was mixed with ammonium sulfate slowly to the concentration of 60%, and then mixed with 3ml cooling acetone (refrigeration at -18 °C for 12 hours) slowly, after standing for 24 hours, centrifugated at 5000rpm for 10 minutes at 4°C. Material at the interface was collected and air-dried as higher purity Grifolan Synthase. The higher purity Grifolan Synthase was dispersed in the native-PAGE gel, and the active band of native-PAGE gel was cut down and broken by ultrasonic treatment for 1min, then centrifugated at 5000rpm for 10 minutes at 4°C, the supernatant was taken and mixed with ammonium sulfate slowly to the concentration of 60%, after 24 hours on standing, centrifugated at 12000rpm for 10 minutes at 4°C, the pellets was got and the purity was checked by SDS-PAGE. The molecular weight of Grifolan Synthase was determined by SDS polyacrylamide gel electrophoresis (SDS-PAGE). Results Grifolan Synthase purified by recrystallization and native-PAGE was checked to be a single band by SDS-PAGE. The data of molecular weight obtained by SDS-PAGE showed that the molecular weight of Grifolan Synthase was 55000Da. Conclusion A purification method of Grifolan Synthase crude extract from the mycelium of Grifola frondosa was researched out, and the molecular weight of grifolan synthase was studied in this paper. It can lay the foundation for the further study on the structure and function of Grifolan Synthase.

scholarly journals ANTIOXIDANT ACTIVITY OF LECTIN ISOLATED FROM LEAVES OF BRYOPHYLLUM PINNATUM

2021 ◽  
Vol 21 (no 1) ◽  
Author(s):  
Sruchi Devi ◽  
Sourbh Suren Garg ◽  
Arvind Kumar ◽  
Nitish Kaushal
Keyword(s):  
Molecular Weight ◽  
Antioxidant Activity ◽  
Ammonium Sulfate ◽  
Normal Saline ◽  
Protein Concentration ◽  
Sds Page ◽  
The Family ◽  
Bryophyllum Pinnatum

The Bryophyllum pinnatum refers to the family Crassulaceae. The glycoprotein from the 0.9% of normal saline extracts of Bryophyllum pinnatum leaves was refined by dialysis with 50% of ammonium sulfate. Protein concentration was invented by Lowry’s method. The dialyzed sample allowed for SDS-PAGE to determine the molecular weight (Mr ). The antioxidant activity of crude lectin extract has been assessed by the DPPH (1, 1-Diphenyl-2-picrylhydrazyl) and ABTS methods. The present study aims to verify the antioxidant activity of lectins detached from the leaves of Bryophyllum pinnatum.

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