Studies on Protein Extraction from Caragana korshinskii Kom. Using Acetone Precipitation Method

2013 ◽  
pp. 1609-1616
Author(s):  
Cheng Zhong ◽  
Zhao Zhou ◽  
Rui Wang ◽  
Peipei Han ◽  
Shiru Jia
Keyword(s):  
Protein Extraction ◽  
Precipitation Method ◽  
Acetone Precipitation ◽  
Caragana Korshinskii

scholarly journals Modified TCA/acetone precipitation of plant proteins for proteomic analysis

2018 ◽  
Author(s):  
Liangjie Niu ◽  
Hang Zhang ◽  
Zhaokun Wu ◽  
Yibo Wang ◽  
Hui Liu ◽  
...  
Keyword(s):  
Proteomic Analysis ◽  
Protein Modification ◽  
Protein Extraction ◽  
Extended Period ◽  
Extraction Methods ◽  
Precipitation Method ◽  
Plant Proteins ◽  
Prolonged Incubation ◽  
Acetone Precipitation ◽  
Modified Method

AbstractProtein extracts obtained from cells or tissues often require removal of interfering substances for the preparation of high-quality protein samples in proteomic analysis. A number of protein extraction methods have been applied to various biological samples. TCA/acetone precipitation and phenol extraction, a common method of protein extraction, is thought to minimize protein degradation and activity of proteases as well as reduce contaminants like salts and polyphenols. However, the TCA/acetone precipitation method relies on the complete pulverization and repeated rinsing of tissue powder to remove the interfering substances, which is laborious and time-consuming. In addition, by prolonged incubation in TCA/acetone, the precipitated proteins are more difficult to re-dissolve. We have described a modified method of TCA/acetone precipitation of plant proteins for proteomic analysis. Proteins of cells or tissues were extracted using SDS-containing buffer, precipitated with equal volume of 20% TCA/acetone, and washed with acetone. Compared to classical TCA/acetone precipitation and simple acetone precipitation, this protocol generates comparable yields, spot numbers, and proteome profiling, but takes less time (ca. 45 min), thus avoiding excess protein modification and degradation after extended-period incubation in TCA/acetone or acetone. The modified TCA/acetone precipitation method is simple, fast, and suitable for proteomic analysis of various plant tissues in proteomic analysis.

Developing a High Efficient Process Integrating Protein Extraction with Cellulosic Ethanol Production from Caragana korshinskii Kom

2012 ◽  
Vol 518-523 ◽  
pp. 5545-5549
Author(s):  
Cheng Zhong ◽  
Rui Wang ◽  
Li Hua Fan ◽  
Shi Ru Jia ◽  
Zhao Zhou
Keyword(s):  
Ethanol Production ◽  
Cellulosic Ethanol ◽  
Protein Extraction ◽  
Cellulose Degradation ◽  
Whole Process ◽  
Caragana Korshinskii ◽  
Useful Components ◽  
High Efficient ◽  
Alkali Hydrolysis ◽  
Pretreated Biomass

Caragana Korshinskii Kom is a widely grown deciduous shrub in the region of inner magnolia. A full utilization of its useful components holds great potential for cellulosic ethanol production in an efficient and economic way. The remaining residues after protein extraction contain 29.02% cellulose and 19.61% hemicellulose, offers a potential feedstock for cellulosic ethanol production. Fermentable capability of hydrolysate derived from pretreated biomass is strongly dependent on pretreatment methods. In this work, three hydrolysis methods (alkaline hydrolysis, acid hydrolysis and acid-alkali hydrolysis) were compared by its digestibility. Alkali pretreatment could help greatly to cellulose degradation. The conversion percentage of cellulose and hemicellulose was increased to 1.43% and 13.01%, respectively. Without any nutrient addition, hydrolysate from alkaline pretreated biomass showed relative fermentable capability by Candida Tropicalis with a final ethanol concentration of 5.4 g/L. A mass balance of the whole process integrating protein extraction and cellulosic ethanol production indicated that 1000kg C. Korshinskii Kom leaves will produce 154 kg crude proteins, and the residues will produce 18 kg ethanol.

scholarly journals Evaluation of female Aedes aegypti proteome via LC-ESI-MS/MS using two protein extraction methods

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e10863
Author(s):  
Abubakar Shettima ◽  
Intan H. Ishak ◽  
Syahirah Hanisah Abdul Rais ◽  
Hadura Abu Hasan ◽  
Nurulhasanah Othman
Keyword(s):  
Extraction Method ◽  
Protein Identification ◽  
Protein Extraction ◽  
Adaptor Protein ◽  
Control Measures ◽  
Protein Yield ◽  
Mass Spectrometry Analysis ◽  
Functional Protein ◽  
Acetone Precipitation ◽  
Esi Ms

Background Proteomic analyses have broadened the horizons of vector control measures by identifying proteins associated with different biological and physiological processes and give further insight into the mosquitoes’ biology, mechanism of insecticide resistance and pathogens-mosquitoes interaction. Female Ae. aegypti ingests human blood to acquire the requisite nutrients to make eggs. During blood ingestion, female mosquitoes transmit different pathogens. Therefore, this study aimed to determine the best protein extraction method for mass spectrometry analysis which will allow a better proteome profiling for female mosquitoes. Methods In this present study, two protein extractions methods were performed to analyze female Ae. aegyti proteome, via TCA acetone precipitation extraction method and a commercial protein extraction reagent CytoBusterTM. Then, protein identification was performed by LC-ESI-MS/MS and followed by functional protein annotation analysis. Results The CytoBusterTM reagent gave the highest protein yield with a mean of 475.90 µg compared to TCA acetone precipitation extraction showed 283.15 µg mean of protein. LC-ESI-MS/MS identified 1,290 and 890 proteins from the CytoBusterTM reagent and TCA acetone precipitation, respectively. When comparing the protein class categories in both methods, there were three additional categories for proteins identified using CytoBusterTM reagent. The proteins were related to scaffold/adaptor protein (PC00226), protein binding activity modulator (PC00095) and intercellular signal molecule (PC00207). In conclusion, the CytoBusterTM protein extraction reagent showed a better performance for the extraction of proteins in term of the protein yield, proteome coverage and extraction speed.

scholarly journals Comparative Analysis of Universal Protein Extraction Methodologies for Screening of Lipase Activity from Agricultural Products

Catalysts ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 816
Author(s):  
Jisu Ha ◽  
Jun-Young Park ◽  
Yoonseok Choi ◽  
Pahn-Shick Chang ◽  
Kyung-Min Park
Keyword(s):  
Lipase Activity ◽  
High Throughput Screening ◽  
Protein Extraction ◽  
Extraction Yield ◽  
Agricultural Products ◽  
Acetone Precipitation ◽  
Commercial Kits ◽  
Protein Assays ◽  
Plant Sources ◽  
Universal Protein

Protein extraction techniques are absolutely required for the research of biological catalysts. The present study compared four universal protein extraction methodologies (ammonium sulfate precipitation, TCA/acetone precipitation, and two commercial kits) to provide practical information on protein extraction in order to discover a novel lipase in agricultural products. Yields of protein extraction from 24 domestic agricultural products and their specific activities were evaluated and compared with each other. TCA/acetone precipitation showed a relatively higher extraction yield (on average, 3.41 ± 1.08 mg protein/0.1 g sample) in crude protein extraction, whereas the Pierce™ Plant Total Protein Extraction Kit showed the highest specific lipase activity on average in both spectrophotometric (266.61 ± 235.78 μU/mg protein) and fluorometric (41.52 ± 32.63 μU/mg protein) assays. Our results suggest that commercial kits for the rapid extraction of soluble functional proteins would be a better choice than conventional precipitation techniques to perform the high-throughput screening of enzyme activity from plant sources. Finally, several agricultural products such as cordyceps, pepper, bracken, and hemp, all of which exhibited an excellent specific lipase activity, were proposed as promising candidates for a source of novel lipases.

scholarly journals Optimizing Human Bile Preparation for Two-Dimensional Gel Electrophoresis

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Hao-Tsai Cheng ◽  
Sen-Yung Hsieh ◽  
Chang-Mu Sung ◽  
Betty Chien-Jung Pai ◽  
Nai-Jen Liu ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Protein Extraction ◽  
Protein Profile ◽  
Two Dimensional Gel Electrophoresis ◽  
Protein Preparation ◽  
Preparation Methods ◽  
Human Bile ◽  
Acetone Precipitation ◽  
Exogenous Compounds ◽  
Dimensional Electrophoresis

Aims. Bile is an important body fluid which assists in the digestion of fat and excretion of endogenous and exogenous compounds. In the present study, an improved sample preparation for human bile was established.Methods and Material. The method involved acetone precipitation followed by protein extraction using commercially available 2D Clean-Up kit. The effectiveness was evaluated by 2-dimensional electrophoresis (2DE) profiling quality, including number of protein spots and spot distribution.Results. The total protein of bile fluid in benign biliary disorders was 0.797 ± 0.465 μg/μL. The sample preparation method using acetone precipitation first followed by 2D Clean-Up kit protein extraction resulted in better quality of 2DE gel images in terms of resolution as compared with other sample preparation methods. Using this protocol, we obtained approximately 558 protein spots on the gel images and with better protein spots presentation of haptoglobin, serum albumin, serotransferrin, and transthyretin.Conclusions. Protein samples of bile prepared using acetone precipitation followed by 2D Clean-Up kit exhibited high protein resolution and significant protein profile. This optimized protein preparation protocol can effectively concentrate bile proteins, remove abundant proteins and debris, and yield clear presentation of nonabundant proteins and its isoforms on 2-dimensional electrophoresis gel images.

scholarly journals Trichloroacetic Acid/Acetone Precipitation Method to Optimize Canine Synovial Fluid for One and Two-Dimensional Gel Electrophoresis Studies

2021 ◽  
Vol 50 (11) ◽  
pp. 3297-3302
Author(s):  
Wei Miao Tan ◽  
Seng Fong Lau ◽  
Sharifah Zakiah Syed Sulaiman ◽  
Nor Shahira Solehah Umran ◽  
Mokrish Ajat ◽  
...  
Keyword(s):  
Synovial Fluid ◽  
Gel Electrophoresis ◽  
Trichloroacetic Acid ◽  
Cost Effective ◽  
Precipitation Method ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Acetone Precipitation ◽  
Cost Effective Technique ◽  
Cost Efficient

The challenge associated with the use of synovial fluid for osteoarthritic proteome studies is the optimization step, which involves the depletion of high abundant proteins from the samples. The objective of this study was to develop a cost efficient and effective method to remove albumin from canine synovial fluid for proteome studies. Pooled synovial fluid samples were obtained from clinically healthy dogs (n=5), with no radiographic features of osteoarthritis. The acetone precipitation method and 10% w/v of trichloroacetic acid/acetone were chosen to deplete the albumin from canine synovial fluid and the outcome from the different methods were compared using one dimensional and two-dimensional gel electrophoresis studies. The results showed that the 10% w/v TCA/acetone precipitation method removed highly abundant proteins from synovial fluid for gel electrophoresis studies compared to the acetone precipitation method. The 10% w/v TCA/acetone precipitation method provides an effective method to remove albumin from the synovial fluid using gel electrophoresis, especially two-dimensional gel electrophoresis. The accessibility and cost of TCA and acetone make this method of precipitation a simple and cost-effective technique in preparing a canine synovial fluid for two-dimensional gel electrophoresis analysis.

scholarly journals Evaluation of protein extraction methods to obtain protein concentrate from cassava leaf

2013 ◽  
Vol 33 (6) ◽  
pp. 1223-1233 ◽  
Author(s):  
Priscila F. Coldebella ◽  
Simone D. Gomes ◽  
Janete A. Evarini ◽  
Marney P. Cereda ◽  
Sílvia R. M. Coelho ◽  
...  
Keyword(s):  
Protein Extraction ◽  
Extraction Methods ◽  
Extraction Yield ◽  
Precipitation Method ◽  
Isoelectric Precipitation ◽  
Manihot Esculenta Crantz ◽  
High Fiber ◽  
Cassava Leaves ◽  
Alternative Protein ◽  
Extraction Processes

The cassava leaf, waste generated in the harvest of the roots, is characterized by high content of protein, vitamins and minerals; however, its use is limited due to the high fiber content and antinutritional substances, which can be removed by obtaining protein concentrates. In this context, the objective of this study was to evaluate protein extraction processes, aiming the use of cassava leaves (Manihot esculenta Crantz) as an alternative protein. Four methods were tested: 1) Coagulation of Proteins by Lowering the Temperature, 2) Extraction by Isoelectric Precipitation, 3) Solubilization of Proteins and 4) Fermentation of Filter Leaf Juice. To obtain the concentrates, the use of fresh or dried leaves and extraction in one or two steps were also evaluated. The solubilization of proteins (method 3) showed a higher extraction yield; however, with concentrate of low quality. The fermentation of the juice (method 4) produced concentrates with higher quality and lower costs and the isoelectric precipitation (method 2) promoted the obtention of concentrates in less time, both with good prospects for use. The use of two extraction steps was not advantageous to the process and there was no difference between the use of fresh or dried leaf, and the use of fresh leaves is presented as a good option for the simplicity of the method.

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