Isolation, Purification and Cryopreservation of Cells from Neonatal Bovine Testis

The development and application of spermatogonial stem cell technology have an important significance in animal cloning, preservation of endangered species and spermatogenesis research. In this study, the seminiferous epithlium cells were isolated and purified, the ceils were cryopreserved after identification, and the effects of different purification and cyopreservation methods on bovine testicular cells were studied. The results showed that there were spermatogonial stem cells and sertoli cells in the neonatal bovine seminiferous tubules, differential adherent selection methods could effectively separate these two cell types. Spermatogonial stem cells were positive after AKP, C-kit, and OCT-4 identification; sertoli ceils were positive after oil red O and vimentin identification. Frozen stock solution supplemented with 10% DMSO had the best effect in spermatogonial stem cell cryopreservation, while fxozen stock solution supplemented with 10% of ethylene glycol and 0.1 mmol. L^-1 trehalose had the best effect in sertoli cells crvooreservation.

Isolation and Preparative Purification for Ginkgolides A and B

In this paper a simple preparative method for isolation and purification of ginkgolides A and B was developed,As starting material,a commercially available standardized ginkgo extract (EGb761,containing 24% flavonoid and 6% terpene trilactones) was used,After a pretreatment step,optimized by the uniform design method ,the concentrated intermediate extract with high content of GA and gb(+90%) was separated into the individual terpenes by preparative liquid chromatography eluted with petroleum ether-ethylacetate,Analysis of products was carried out by means of HPLC-ELSD(evaporative light -scattering detector),The results show that ginkgolides A and B are obtained in higher yield and better purity.

Isolation and Purification of Unstable Iridoid Glucosides from Traditional Chinese Medicine by Preparative High Performance Liquid Chromatography Coupled with Solid-phase Extraction

An efficient preparative method was successfully developed for isolation and purification of unstable components from medicinal plant extracts, using a combined method of preparative high performance liquid chro-matography(HPLC) and solid-phase extraction(SPE). The aim of this study was to obtain an effective method with high preparative efficiency and importantly to avoid the transformation of unstable compounds. The preparative HPLC system was based on an LC/MS controlled four-channel autopurification system. The SPE method was performed with a C18 packing material to trap the target compounds and to remove the acidic additive derived from the mobile phase. Using this method, the unstable iridoid glucosides(IGs) as model compounds were successfully isolated and purified from the extract of Hedyotis diffusa Willd. Six IGs(including one new minor IG) and one nucleotide compound were simultaneously obtained, each with a purity of 91% as determined by HPLC. The structures of the isolated compounds were identified by UPLC/Q-TOF MS, UV, 1D and/or 2D NMR. It was demonstrated that the combination of preparative HPLC with SPE is a versatile tool for preparative purification of unstable compounds from complex natural products.

Isolation and Purification of Bovine Colostrum sIgA and IgG

To further utilize bioactive substance such as bovine colostrum sIgA and IgG, sIgA and IgG were isolated and purified simultaneously by salting out, ultrafiltration and gel chromatography, etc. The analysis of results were showed quantitatively by nonhydrogenized SDS-PAGE, and quanlities of sIgA and IgG were respectively detected by Western Blot. The results showed that the purity and yield of bovine colostrum sIgA were 85.3% and 42.8%, respectively, while the purity and yield of bovine colostrum IgG were respectively 97.2% and 64.4%. This preparative method provides theoretical and experimental foundation for sIgA industrial production.

ISOLATION AND PURIFICATION OF METHANE IVf ONOOXYGENASE FROM METHYLOMONAS SPECIES GYJ3

A three—component enzyme system that catalyzes in vivo the oxidation of CH_4 to CH_3OH has been purified with high specific activity from an unusual type I methanotroph through the use of stabilizing reagents.

A Review on the Isolation and Purification of D-allulose

D-allulose has very little content in nature,and it needs to be synthesized artificially and meet the purity requirements of industrial grade.The basic physical and chemical properties of D-allulose,its preparation methods and many different ways of isolation and purification were described.In order to achieve the goal of industrial production of D-allulose as soon as possible,the research progress of D-allulose isolation and purification technologies at home and abroad in recent years was classified and discussed,so as to provide useful reference for the practical improvement of D-allulose isolation and purification process technologies.

Apoptosis and oxidative injury of donor islets during isolation and purification

Objective To observe the changes of islet cell apoptosis and oxidation - antioxidation before transplantation,and to explore pathways of islet protection. Methods Fifteen human pancreases were perfused with Hanks solution containing collagenase,then digested and isolated.

Study on Purification and Properties of Antibacterial Substance from Antagonistic Bacteria P72 against Wheat Scab

[Objective] The study aimed to explore the antibacterial activity and stability of antagonistic bacteria P72 against wheat scab.[Method] The Bacillus subtilis P72 was inoculated into fermentation media with the inoculation amount 5%,then cultured on shaking table at 28 ℃ for 48 h and centrifuged at 8 000 r/min for 10 min.The supernatant of fermentation broth was purified and then the genetic stability,thermal stability and pH stability were detected.[Result] By DEAE-52 ion exchange chromatography,the protein eluted by 0.5 mol/L NaCl solution possessed the strongest antagonistic ability against wheat scab pathogen.The SDS-PAGE electrophoresis showed that the molecular weight of the purified protein with antibacterial effect was 40 kD.By stability test,the antibacterial substance produced by P72 strain showed heritable antagonistic activity,high stability below 60 ℃,stability to acid and unstability to alkali.[Conclusion] The antagonistic bacteria P72 had strong antagonistic ability to wheat scab pathogen,stable antibacterial activity and thermal stability,so it would possess a wide development prospect.

Isolation of Plasma and Thylakoid Membranes from the Heterocystous Cyanobacterium Anabaena sp. PCC 7120

利用水溶性多聚体双相法分离蓝细菌Anabaenasp .PCC 712 0质膜和类囊体膜两种膜系统。吸收光谱分析表明 ,质膜相和类囊体膜相的主要色素分别为类胡萝卜素和叶绿素。SDS_凝胶电泳显示这两种膜系统蛋白组成有很大差别。这种分离方法容易操作 ,对研究蓝细菌的膜蛋白和膜脂非常有用。

Study on purification and ultrastructure of a baculovirus in Penaeus chinensis

A kind of baculovirus was isolated from the cephalothorax homogenate of sick or morbid Penaeus chinensis by differential centrifugation and density gradient centrifugation. Electron microscopic examination of ultrathin section of the gills, stomach and mid-gut tissues also revealed the presence of rod-shaped baculoviral particles with the same size in the affected cell nuclei, where most of the virions arranging in cluster assembled and caused a series of cytopathic changes. The virion covered with bilaminal envelope was 320 ~ 400 nm × 100 ~ 130 nm in size, whereas the nucleocapsid ranged in size of 250~ 300 nm in length and 70 ~ 100 nm in breadth respectively. No nuclear polyhedron or granulin occlusion theies have been found in cells. According to the principle of viral classification, this newly found virus could probably belong to the non-occluded subgroup of insect baculoviridae, i. e., C subgroup baculovirus.

Purification and Characterization of a Novel Lipase from Antarctic Krill

Lipase from Antarctic krill,with a molecular weight of 71.27kDa,was purified with ammonium sulfate precipitation and a series of chromatographic separations over ion exchange(DEAE)and gel filtration columns(Sephacryl S-100),resulting in 5.2%recovery with a 22.4-fold purification ratio.The optimal pH and temperature for enzyme activity were 8.0 and 45℃,respectively.Purified lipase had Km and Vmax values of 3.27mmolL−1 and 2.4Umg−1,respectively,using p-nitrophenyl laurate as the substrate.Lipase activity was enhanced by adding Ca2+and Mg2+ions in the concentration ranges of 0–0.5mmolL−1 and 0–0.3mmolL−1,respectively,while the activity was inhibited by a further increase in these ion concentrations.Fe3+and Cu2+ions showed obvious inhibitory effects on enzyme activity,and the inhibition rates were 71.8%and 53.3%when the ion concentrations were 0.5mmolL−1.

Isolation and identification of chemical constituents from the vinegar-prepared Corydalis yanhusuo

Phytochemical investigation of the vinegar-prepared Corydalis yanhusuo led to the isolation of one aristolactam derivative,1,2,8,9-tetramethoxy-5-methyldibenzo[cd,f]indol-4-(5H)-one(1),and seven aporphine alkaloids,including 2,9,10-trimethoxydibenz[de,g]quinolin-7-one(2),1-hydroxy-2,9,10-trimethoxy-7H-dibenzo(de,g)quinoline-7-one(3),oxoglaucine(4),N-methyloxoglaucine trifluoroacetate trifluoroacetate(5),corunine acetate(6),pontevedrine(7),and oxoglaucidaline trifluoroacetate(8).The structures of the isolated compounds were elucidated by extensive spectroscopic data analysis and comparison with the previous reports.Among them,compounds 1 and 2 were obtained as natural products for the first time,and their NMR data were unambiguously assigned.In addition,compound 1 exhibited moderate cytotoxic activity against HepG2 cells with an IC50 value of 16.0±6.4μM.

Purification and Partial Characterization of an Antibacterial Protein Tz1

A strain (T3) of Bacillus has been screened from paddy field. It secretes large amount of antibacterialproteins which show a strong inhibiting activity against several pathogens of rice. This paper presentsa systematic study of the inhibition spectrum and characteristics of T3 proteins. Total proteins wereprecipitated with ammonium sulfate at 70% saturation from cell-free culture. One of the proteins(Tzl) was purified from the crude extracts with Sephadex G-100, DEAE52 and FPLC Superose 12columns. A single band was demonstrated in both 15% SDS-PAGE and IEF, with an apparent MWof 6,9 kd and a pI of 7.8. Its amino acid composition was analyzed and part of its sequence,determined.

Study on Extraction and Separation of Dihydromyricetin from Vine Tea [Ampelopsis grossedentata(Hand-Mazz) W. T. wang]

Ampelopsis grossedentata(Hand-Mazz) W. T. wang is a typical edible plant with important physiological health functions. Dihydromyricetin is the main active ingredient of A. grossedentata, which has good pharmacological effects such as antibacterial, anti-inflammatory, anti-oxidation, anti-tumor and liver protecting effects. This paper summarized the research progress on the extraction materials, extraction methods and separation and purification processes of dihydromyricetin in A. grossedentata to understand the best experimental method of dihydromyricetin, aiming to provide an important theoretical basis for the comprehensive development and utilization of dihydromyricetin.

Preparation and antiviral effects of polysaccharides from Panax japonicus

Water-soluble polysaccharides were prepared from Panax japonicus by hot water extraction and ethanol precipitation.The polysaccharides were further purified by ion exchange chromatography to obtain neutral and acidic polysaccharides.The neutral polysaccharide fraction mainly contained Glc(90.2%),which was a glucan fraction.The acidic polysaccharide fraction mainly contained GalA(43.6%),Gal(21.7%),and Ara(15.4%),with a degree of methyl-esterification of 20.3%,which was a pectic polysaccharide.The acidic polysaccharide of Panax japonicus could effectively inhibit the replication of human seasonal influenza virus H1N1 and canine influenza virus H3N2 in MDCK cells and A549 cells and significantly reduce the virus titer in infected cells.It also effectively inhibited the number of infected cells of the SARS-CoV-2 South Africa strain and the Omicron strain.The acid polysaccharide of Panax japonicus showed good efficacy against influenza virus and COVID-19 infection,which could be used as a potential antiviral candidate drug molecule in the future.

Isolation and Structural Analysis of an Acidic Polysaccharide from Astragalus membranaceus （Fisch.） Bunge

A new water-soluble hetero-polysaccharlde, APSID3, was obtained from a hot-water extract of the roots of Astragalus membranaceus （Flsch.） Bunge by DEAE-Sepharose Fast Flow and Sephacryl S-300 chromatography. The molecular weight of APSID3 was estimated to be 5.79 × 10^5 Da. Based on a sugar composlUon analysis, methylatlon analysis, partial hydrolysis and 13C nuclear magnetic resonance experimentation, It was concluded that the minimal repeat unit of APSID3 was composed of one terminal arablnose, one 1,5-1Inked arabinose, one 1,3-1Inked rhamnose, one 1,3,4-1Inked rhamnose, five 1,4-1Inked methyl galacturonates and six 1,4-1inked methyl glucuronates.

In vitro Study on Human Trophoblast Cells Infected with HCMV

Human trophoblast cells were isolated and cultured in vitro in order to investigate possible pathogenesis of intrauterine infection caused by HCMV.Trophoblast cells were obtained by compound enzymes digestion and discontinuous percoll gradient.Cells and purity were identified by using immunocytochemistry assay with anti-CK7,Vim and β-hCG antibodies.HCMV AD169 strain replication in isolated trophoblast cells and cell apoptosis were detected at different time points post infection(p.i.).The results showed tha...

Bacterial extracellular vesicles as bioactive nanocarriers for drug delivery:Advances and perspectives

Nanosized extracellular vesicles derived from bacteria contain diverse cargo and transfer intercellular bioactive molecules to cells.Due to their favorable intercellular interactions,cell membrane-derived bacterial extracellular vesicles(BEVs)have great potential to become novel drug delivery platforms.In this review,we summarize the biogenesis mechanism and compositions of various BEVs.In addition,an overview of effective isolation and purification techniques of BEVs is provided.In particular,we focus on the application of BEVs as bioactive nanocarriers for drug delivery.Finally,we summarize the advances and challenges of BEVs after providing a comprehensive discussion in each section.We believe that a deeper understanding of BEVs will open new avenues for their exploitation in drug delivery applications.

Three new ursane-type triterpenoids from Rosmarinus officinalis and their biological activities

Three new ursane-type triterpenoids,3-oxours-12-en-20,28-olide(1),3β-hydroxyurs-12-en-20,28-olide(2)and 3β-hydroxyurs-11,13(18)-dien-20,28-olide(3),were isolated from a potent anti-inflammatory and antibacterial fraction of the ethanolic extract of Rosmarinus officinalis.Their structures were elucidated by a combination of extensive 1D-and 2D-NMR experiments,MS data and comparisons with literature reports.Compounds 1-3 exhibited significantly inhibitory effects on nitric oxide production in lipopolysaccharide-activated mouse RAW264.7 macrophages,but no antibacterial activity was found at a concentration of 128μg·mL^(-1).