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Chromatographic fractions of a rough extract of echinoid spawn (REES) is demonstrated to efficiently induce oocyte maturation in aspidochirote holothuroids. The method is so efficient that it is currently used in holothuriculture to obtain fertilised eggs even outside the reproductive period of the aquacultivated species. In the present study, we isolated and identified from echinoid spawns the molecule responsible for the induction of the oocyte maturation in the Mediterranean holothuroid Holothuria tubulosa and the Indo-Pacific Holothuria scabra. The use of proteinase K and dialysing membranes suggests that the active molecule is a protein with a molecular weight superior to 12,000 kDa. The active molecule has been isolated on G-100 Sephadex chromatography column. Active chromatographic fractions include seven proteins identified with nanoLC-MS/MS technique. One of them, identified as a thioredoxin-2 and to which the name Trx-REES was given, gave levels of maturation similar to those obtained with the REES and with a commercial thioredoxin extracted from Escherichia coli. Occurrence of thioredoxin in REES was confirmed by immunoblot analysis, and the maturation-inducing properties of thioredoxin were positively checked using anti-thioredoxin antibodies. A peptide of 6 AAs corresponding to the active site of Trx-REES, composed of WCNPCK, was synthesised and its efficiency in holothuroid oocyte maturation was tested. At some concentrations, the isolated peptide was 1.2 times more active than the REES. This short peptide may be used as an effective tool for artificial induction of oocyte maturation in the sea cucumber aquaculture context.

Bacillus velezensis BS2 was isolated from meongge (common sea squirt) jeotgal, a Korean fermented seafood, and produces a bacteriocin, BacBS2, which strongly inhibits Listeria monocytogenes and Bacillus cereus. BacBS2 was partially purified by Q-Sepharose column chromatography after ammonium sulfate precipitation of the culture supernatant, then further purified by Sephadex G-50 column chromatography. Partially purified BacBS2 was estimated to be 6.5 kDa in size by Tricine-SDS PAGE and activity detection by gel-overlay. Enzyme treatment and FT-IR spectrum of partially purified BacBS2 confirmed its proteinaceous nature. BacBS2 was fully stable at pH 4-9, and half of activity was retained at pH 1-3. Full activity was retained after exposure to 80°C for 15 min, but half of the activity was retained upon exposure to 90°C for 15 min or 100°C for 10 min. BacBS2 inhibited L. monocytogenes by bactericidal mode of action. B. velezensis BS2 and its BacBS2 seem useful as biopreservatives for fermented foods such as jeotgal.

The study is devoted to clarifying the regulatory and catalytic properties of glutathione peroxidase from liver of rats with toxic liver damage and 6-hydroxy-2,2,4-trimethyl-1,2-dihydroquinoline exposure. The pathology was induced by intragastric administration of carbon tetrachloride at a dose of 0.064 cm3 per 100 g of body weight. The test compound was orally administered at a dose of 50 mg/kg., Animals were divided into 3 groupsduring the experiment: control rats, animals with toxic liver damage, and rats receiving 6-hydroxy-2,2,4-trimethyl-1,2-dihydroquinoline against the background of the pathology. For the purification of glutathione peroxidase from rat liver, chromatographic separation methods with Sephadex G-25 and DEAE-cellulose were used. Glutathione peroxidase activity was measured by a conjugate reaction with glutathione reductase using the Warburg test. The protein content was assessed using the BCA protein assay kit. The results showed that in rats treated with 6-hydroxy-2,2,4-trimethyl-1,2-dihydroquinoline against the background of the pathology induction, the pH optimum and glutathione peroxidase affinity for glutathione were changed in the controldirection. The obtained data may indicate a change in the conformational properties of the enzyme under conditions of a decrease the free radical oxidation intensity under the test compoundaction.In addition, the introduction of 6-hydroxy-2,2,4-trimethyl-1,2-dihydroquinoline contributed to the normalization of the glutathione peroxidase regulatory properties, to respect toisocitrate, glucose-6-phosphate and Fe2+. Thus, the results of the work indicate the ability of 6-hydroxy-2,2,4-trimethyl-1,2-dihydroquinoline to have a corrective effect on the glutathione peroxidase catalytic and regulatory properties due, apparently, to the antioxidant properties manifestation by this compound.

A cellulase from the extreme obligate halophilic fungus, Aspergillus flavus, isolated from a man-made solar saltern in Phetchaburi, Thailand, was purified by ammonium sulfate precipitation and using Sephadex G-100 gel filtration column chromatography. The cellulase was found to be approximately 55 kDa by SDS-PAGE. Using CMC as a substrate, the specific activity of the cellulase was 62.9 units (U) mg−1 with Vmax and Km values of 37.87 mol min−1 mg−1 and 3.02 mg mL−1, respectively. Characterization of the enzyme revealed it to be an extremozyme, having an optimum activity at pH 10, 60 °C, and 200 g L−1 of NaCl. The enzyme activity was not significantly altered by the addition of divalent metal cations at 2 mM and neither did s-mercaptoethanol, while EDTA was found strongly inhibiting the cellulase. Compared with commercial cellulase, the purified cellulase from A. flavus was more active in the extremity of conditions, especially at pH 10, 60 °C, and 150 g L−1 NaCl, whereas the commercial cellulase had a very low activity.

Angiotensin-converting enzyme inhibitory peptides were isolated and identified from milk fermented using Lactobacillus delbrueckii QS306. The peptide with the highest angiotensin-converting enzyme inhibitory activity (C5) was purified using ultrafiltration with 10 and 3 kDa molecular mass cut-off membranes, Sephadex G-15 (Sigma-Aldrich, St. Louis, MO) gel filtration chromatography, reversed-phase HPLC, and Orbitrap Elite (Thermo Fisher Scientific Inc., Waltham, MA) liquid chromatography-tandem mass spectrometry. We obtained peptide LPYPY by microbial fermentation, which was derived from κ-casein f (AA 77–81). We synthesized LPYPY using an Fmoc solid-phase synthesis method and explored the secondary structure of the pentapeptide. The half maximal inhibitory concentration for the angiotensin-converting enzyme inhibitory activity of LPYPY was 12.87 μg/mL. The results provide additional information for ongoing research and the development of functional foods having antihypertensive effects.

The preparation and fractionation of oligomeric proanthocyanidins (OPCs) are particularly important for the application of tannins in the biomedical field. By use of two different methods-gel filtration chromatography (GFC) with Sephadex LH-20 and progressive solvent precipitation-the OPCs were prepared and fractionated from mangosteen pericarp. The fractions were compared by reversed-phase and normal-phase high-performance liquid chromatography-electrospray ionization mass spectrometry and gel permeation chromatography. GFC directly purified oligomers (monomer to pentamer) with polydispersity values close to 1 and generated fractions with a higher level of total phenols (800.59 mg gallic acid equivalents per gram) but a lower yield (7.72%). Progressive solvent precipitation rapidly prepared and fractionated OPCs with a lower level of total phenols (609.57 mg gallic acid equivalents per gram) but a higher yield (24.74%) and higher polydispersity. Additionally, we found pronounced structural and quantitative differences among different tannin-rich fractions, and fractions obtained by GFC better reflected the structural diversity and complexity of OPCs from mangosteen pericarp. This study presents different ways of preparing and fractionating OPCs in the biomedical field.

Isolation and characterization of a bacteriocin from Brevibacillus laterosporus DS-3 and its ability in producing antibacterial activity were examined in this study. We isolated 48 bacterial strains from different native ecological niches. Sources included lake water, soil, air, and food. Among the isolates from the soil sample, morphological and biochemical tests identified Brev. laterosporus DS-3, which showed great potential for bacteriocin production. It was purified by ammonium sulfate precipitation, then applied in a Sephadex G-15 gel filtration column connected to a purification of AKTA Pure (USA) and reverse phase chromatography. MALDI-TOF mass spectrometry indicated that bacteriocin DS-3 has a low molecular weight of 1592.579 Da. LNTLETEEWFFK was identified as the sequence by mass spectrometry analysis in tandem liquid chromatography and MASCOT research. This bacteriocin inhibited Gram-positive and Gram-negative bacteria and was effective against Methicillin-resistant Staphylococcus aureus (MRSA). Brev. laterosporus DS-3 present in here is thermostable and tolerant to extreme pH. However, inactivation of its antibacterial activity was observed when treated with papain and trypsin but not pepsin, proteinase K, and catalase.

Rituximab is a chimeric monoclonal antibody which has specific for CD20 antigen expressed by pre-B and mature B-cells. Radiolabelled Rituximab, 131 I-Rituximab, has been sucessfully used for treatment of B-Cell NHL. Due to its short shelf-life, 131 I-Rituximab is commonly freshly prepared in hospitals prior to its used. This study aimed to validate rituximab iodination procedure for 131I-Rituximab preparation in order to find the most suitable procedure to be applied in hospitals which intend to produce 131 I-Rituximab in-house. Three different methods of radiolabelling using three types of oxidizing agents, namely Iodobeads, Iodogen, and Chloramine-T were performed. Prior to the validation, radiochemical purity test and purification procedures were also validated as these procedures are critical for producing an acceptable quality of I-Rituximab. In addition, the shelf-life of 131 I-Rituximab was also studied. This study was conducted at the Centre for Radioisotope and Radiopharmaceutical Technology, Serpong during the period of July 2015 to February 2018. The results showed that the radiochemical purity test of 131 I-Rituximab could be easily performed by using instance thin layer chromatography–silica gel (ITLC-SG) in the stationary phase and 85% methanol or saline in the mobile phase. Purification of 131 I-Rituximab was conducted using a Sephadex G-25 M filled column with 0.1 M PBS, pH 7.2, as the eluent that was found to be quite reliable to give 131 I-Rituximab with radiochemical purity of >95% and recovery of approximately 90%. Radiolabelling efficiency performed using Iodobeads was the lowest (60%) compared to that of Iodogen and Chloramine-T (80–90%). In addition, approximately 30% of I was retained by Iodobeads and this procedure was time consuming(~ 1 hours). It is concluded that Chloramine-T and Iodogen are better than Iodobeads as the oxidizing agent for radiolabelling of Rituximab with 131 I. The radiochemical purity of 131 I-Rituximab is well maintained when stored at room temperature and in 4 °C temperature up to 6 hours. Validasi Prosedur Iodinasi Rituximab untuk Preparasi 131 I-Rituximab Validasi proseduri odinasi rituximab untuk preparasi 131 I-Rituximab telah berhasil dilakukan. Validasi ini dilakukan untuk mendapatkan prosedur yang paling sesuai yang dapat diaplikasikan untuk produksi 131 I-Rituximabdi rumah sakit yang ingin memproduksi 131 I-Rituximab di laboratorium mereka. Tiga metode radiolabelling menggunakan 3 jenis oksidator Iodobeads, Iodogen, dan Chloramine-T telah divalidasi. Sebelum validasi ini, prosedur uji kemurnian radiokimia dan pemurnian divalidasi terlebih dahulu karena prosedur-prosedur ini sangat berpengaruh dalam penyediaan 131 I-Rituximab dengan kualitas yang baik. Disampingitu, lama simpan 131 I-Rituximab juga dipelajari. Penelitan ini dilaksanakan di Pusat Teknologi Radioisotop dan Radiofarmaka, Serpong, Juli 2015–Februari 2018. Hasil penelitian memperlihatkan bahwa uji kemurnian radiokimia 131 I-Rituximab dapat dilakukan dengan mudah menggunakan instance thin layer chromatography – silica gel (ITLC-SG) sebagai fasa diam dan metanol 85% atau larutan salin sebagai fasa gerak. Pemurnian 131 I-Rituximab menggunakan kolom Sephadex G-25 M dan0.1 M PBS pH 7,2 sebagai eluen dapat diandalkan dan memberikan 131 I-Rituximab dengan kemurnian radiokimia >95% dan sekitar 90% perolehan kembali. Efisiensi penandaaan menggunakan Iodobeads didapatkan paling (60%) dibanding dengan Iodogen dan Chloramine -T (80 – 90%). Di samping itu, sekitar 30% 131 I hilang karena terikat pada Iodobeads dan prosedur ini memakan waktu yang panjang (~1 jam). Penandaan Rituximab 131 I menggunakan Chloramine -T and Iodogen dapat disimpulkan lebih baik dibanding dengan menggunakan Iodobeads . Kemurnian radiokimia 131 I-Rituximab terjaga dengan baik pada penyimpanan selama 6 jam pada suhu kamar dan 4 °C.

This study aimed to specifically prepare angiotensin I-converting enzyme (ACE) inhibitory peptides rich in C-terminal proline from oyster proteins using chymotrypsin and proline-specific endopeptidases (PSEases). The hydrolysates were purified with Sephadex G25, Superdex™ 30 Increase 10/300 GL gel filtration chromatography and reverse-phase high-performance liquid chromatography (RP-HPLC). The ACE inhibitory IC50 of purified fraction (G1E1C1) was 0.032 ± 0.003 mg/mL. According to ESI–MS and ESI–MS/MS analyses, there were three novel ACE inhibitory peptides in G1E1C1 fraction. Their sequences were Ser-Ala-Pro, Ala-Met-Pro and Thr-Ser-Gly-Pro. Molecular docking of peptides to ACE was studied. Metal–acceptor interactions and conventional hydrogen bonds greatly promoted the stability of peptides to ACE interaction. Pyrrole ring of proline may lead to higher inhibitory activity of peptides. It easily formed a Pi–alkyl interaction with aromatic ring residues (His353, His387, His513, and Phe512). Pi interactions may promote the effect of peptides on ACE. Also, C atom adjacent to the N atom of the pyrrole ring easily formed a carbon hydrogen bond with Ala354. The research discovered three novel ACE inhibitory peptides and provided a method to obtain ACE inhibitory peptides with specific structure X-Pro. The research result played an important role in revealing the structure–activity relationship of ACE inhibitory peptides and designing novel peptides with enhanced biological activity.

Plant defensins are subgroup of plant antimicrobial peptides that have high potentials in developing safe natural agricultural and pharmaceutical biocontrol products as alternatives for the current chemical pesticides, insecticides, and antibiotics. In this study, the level of defensin gene expression was evaluated in floral buds of 9 native plants from Saudi Arabia environment using reverse transcription and polymerase chain reaction (RT-PCR). As defensins are highly expressed at the mRNA level in Ochradenus baccatus (Ob), it was used for the isolation of defensin peptides using ion exchange chromatography followed by gel filtration using Sephadex G50. SDS-PAGE of ion exchange purified proteins detected several bands in a range of 2–10 kD. These proteins were separated by gel filtration using Sephadex G50. The resultant peptide of 5 kD showed an antimicrobial activity. The peptide inhibited Gram positive and Gram negative bacteria with IC50 ranged from 10 to 25 μg/mL. It also inhibited 25 to 60% of fungal linear growth at concentrations of 12.5 and 25 μg/mL. The present results demonstrate that, we could separate and purify a 5 kD peptide from Ob. This peptide could be utilized to develop natural antimicrobial products.

Thirteen pentacyclic triterpenes, methyl 3-oxours-12-en-23-oate, marsformosanone, taraxerone, β-amyrenone, α-amyrenone, lupenone, 24-methylencycloartan-3-one, moretenol acetate, β-amyrin acetate, germanicol acetate, 24-methylencycloartanyl acetate, β-amyrin, and α-amyrin were identified in a chloroform-methanol propolis extract from Melipona beecheii. Additionally, were identified in this propolis, hexadecanoic acid, methyl ester, octadecanoic acid, methyl ester and 1-triacontanol. The purification of the propolis extract was carried out using different chromatographic techniques, including vacuum liquid chromatography, gravity column chromatography and gel filtration chromatography Sephadex LH-20. The identification of the metabolites was performed using mass spectrometry. Keywords: Stingless bee, Propolis, Pentacyclic triterpenes, β-Amyrin acetate, Germanicol acetate, Lupenone

The influence of faba bean seed fermentation on the release of bioactive peptides after digestion in gastrointestinal conditions was investigated. The molecular mass of protein detected by SDS-PAGE was in the range from 6.5 to 97 kDa. The highest degree of protein hydrolysis (98.72%) was noted for protein obtained from seeds fermented for 3 h at 37 °C. The hydrolyzate of seeds fermented for 7 days at 30 °C was characterized by the highest peptide content (4.78 mg/mL). The highest ACE-inhibitory and antiradical against ABTS·+ activity were noted for a peptide fraction obtained from a sample fermented for 3 days at 30 °C (IC50 = 1.01 and 0.99 mg/mL, respectively). The sample obtained after 3-h fermentation at 22 °C was characterized by the highest LOX inhibitory activity (IC50 = 0.54 mg/mL). The lowest IC50 value was determined for fractions obtained from seeds fermented for 3 h and 7 days at 37 °C, and the value was not significantly different (0.92 and 0.89 mg/mL, respectively). The peptide fraction obtained from seeds fermented for 3 days at 30 °C with the highest potential for inhibition of metabolic syndrome development was separated on Sephadex G10 and peptide sequences were identified by LC-MS/MS.

Ethylenediaminetetraacetic acid (EDTA) and diethylenetriamine-pentaacetic acid (DTPA) are water ‘softening’ agents that are present in numerous household and industrial detergents. Since these particular chelating agents are not significantly degraded during conventional wastewater treatment processes, wastewater treatment plant (WTP) effluents can contain up to 19 µM of EDTA and 7 µM of DTPA. Little, however, is known about the release of EDTA and DTPA from WTPs to rivers. To gain insight, we here report on the development of a cost-effective analytical method. This method is based on the chromatography of a humic acid-cadmium (HA-Cd) complex on a size-exclusion chromatography column (SEC, Sephadex G-15) while using WTP effluents from Lethbridge, Banff and Canmore which contained 10 mM Tris-buffer as the mobile phase (pH 8.2). The intact HA-Cd complex is detected by means of a flame atomic absorption spectrometer (FAAS). The addition of equimolar EDTA and DTPA up to 10 µM allowed us to observe a concentration-dependent increase of the retention time of the main Cd-peak. This behaviour was qualitatively comparable between the WTP effluents and was rationalised by the EDTA/DTPA-mediated mobilisation of Cd from the HA-Cd complex. The signal intensity that corresponded to the mobilised Cd was used to establish calibration curves with corresponding correlation coefficients in the range of 0.950–0.978. Therefore, the developed method yields robust results for realistic concentrations of equimolar EDTA/DTPA in real WTP effluents. The developed method can now be applied to analyse real WTP effluent for the presence of chelating agents, whose concentrations may be expressed as being equivalent to a particular equimolar EDTA/DTPA concentration.

Bacillus circulans 528 produces a restriction endonuclease, Bci528I which is an isoschizomer of EcoRI. We purified the enzyme, using Sephadex G-150, Phospho-cellulose, DEAE-cellulose, Hepharin-Sepharose CL-6B chromatography. The specific activity of Bci528I was 29,400 U/mg·protein. Bci528I recognizes 5'-GAATTC-3' in dsDNA and cleaves between G and A of the recognition sequence, producing a symmetric four base 5'overhang.

Milk whey proteins carry out a number of important biological functions and also they are precursors of many biologically active peptides (antihypertensive peptides, antagonists of opioid receptors, regulators of intestinal motility, immunomodulatory, anti-microbial and anti-cancer peptides, appetite regulators and so on.). An important stage in natural bioactive peptides obtaining from milk whey proteins is the isolation of homogeneous proteins-precursors. Considering the significant difference in the molecular masses of whey proteins, a promising method for their selection is gel filtration. The purpose of the research was the fractionation of bioactive peptides precursors from milk whey using gel filtration on Sephadex G-150. The whey was obtained from fresh skimmed milk after isoelectric precipitation of casein. Gel filtration was carried out on the columns from a liquid chromatography kit by the “Reanal” company. The fractional composition and the degree of homogeneity of milk whey proteins were determined by disc-electrophoresis in the plates of a polyacrylamide gel. A repeated gel filtration of fractions from the chromatographic peaks, separated into sections, was performed to increase the fractionation efficiency. While choosing a dextran gel for gel filtration of precursors of biologically active peptides from milk whey proteins, we have taken into account the range of their molecular weights (from 10000 to 150000 Da), the ability to form supramolecular structures (β-LG), as well as the previously obtained results of gel filtration. As a result, it was shown that repeated gel filtration of milk whey on Sephadex G-150 allows efficiently fractionate the proteins-precursors of bioactive peptides. The range of peptides and proteins molecular weights that can be fractionated on this Sephadex is from 5000 to 300 000 Da. The usage of repeated gel filtration on Sephadex G-150 with the chromatogram separation into sectors allows to effectively fractionate proteins-precursors of bioactive peptides from milk whey. In particular, homogeneous β-lactoglobulin (degree of homogeneity > 95 %) and partially purified α-lactalbumin, as well as a group of immunoglobulins and a proteose-peptone fraction were obtained.

Cranberry ( Vaccinium macrocarpon) juice is traditionally used for the prevention of urinary tract infections. Human urine produced after cranberry juice consumption can prevent Escherichia coli adhesion, but the antiadhesive urinary metabolites responsible have not been conclusively identified. Adult female sows were therefore fed spray-dried cranberry powder (5 g/kg/day), and urine was collected via catheter. Urine fractions were tested for antiadhesion activity using a human red blood cell (A+) anti-hemagglutination assay with uropathogenic P-fimbriated E. coli. Components were isolated from fractions of interest using Sephadex LH-20 gel filtration chromatography followed by HPLC on normal and reversed-phase sorbents with evaporative light scattering detection. Active urine fractions were found to contain a complex series of oligosaccharides but not proanthocyanidins, and a single representative arabinoxyloglucan octasaccharide was isolated in sufficient quantity and purity for full structural characterization by chemical derivatization and NMR spectroscopic methods. Analogous cranberry material contained a similar complex series of arabinoxyloglucan oligosaccharides that exhibited antiadhesion properties in preliminary testing. These results indicate that oligosaccharides structurally related to those found in cranberry may contribute to the antiadhesion properties of urine after cranberry consumption.

Previously reported peptides derived from napin of rapeseed ( Brassica napus) have been shown to inhibit DPP-IV in silico. In the present study, napin extracted from rapeseed was hydrolyzed by commercial enzymes and filtered by an ultrafiltration membrane. The napin hydrolysate was then purified by a Sephadex G-15 gel-filtration column and preparative RP-HPLC. A two-enzyme-combination approach with alcalase and trypsin was the most favorable in terms of the DPP-IV-inhibitory activity (IC50 = 0.68 mg/mL) of the napin hydrolysate. Three peptides and one modified peptide (pyroglutamate mutation at the N-terminus) were identified using HPLC-triple-TOF-MS/MS. DPP-IV-inhibitory activity and the types of enzyme inhibition were also determined. Meanwhile, key residues associated with the interactions between the selected peptides and DPP-IV were investigated by molecular docking. IPQVS has key amino acid residues (Tyr547, Glu205, and Glu206) that are consistent with Diprotin A. ELHQEEPL could form a better covalent bond with Arg358 in the S3 pocket of DPP-IV.

This study investigated the effects of different temperatures on structural characterization and antitumor activity of polysaccharides from Astragalus membranaceus. APS4 and APS90 were extracted at 4°C and 90°C, respectively, and purified by Sephadex G-200 column. APS4-90 were obtained from APS4 after treatment at 90°C for 6h. MTT results showed that APS4 possessed the highest inhibitory effects on MGC-803, A549 and HepG2 cells. HPGPC analysis showed that the average molecular weights of these polysaccharides were approximately 1.5×106Da, while the asymmetrical peak of APS4-90 suggested heat degradation and configuration changes of APS4. GC, NMR and methylation results showed that these three polysaccharides had similar monosaccharide components (mainly contain glucose), and their backbones were composed of (1→2)‑α‑d‑Glcp. However, APS4 showed higher content of (1→2,6)‑α‑d‑Glcp compared to APS4-90 and APS90, which indicated that higher branched degree would be responsible for the stronger in vitro antitumor activity in APS4. These results were also confirmed by specific rotation and SEM analysis. Our study suggested that APS4 had the potential application for cancer treatment.

Lactate dehydrogenase activity (LDH, EC 1.1.1.27) was induced in pea (Pisum sativum L.) leaves by culturing plants under flood conditions. The enzyme was purified to an electrophoretic homogeneous state with a multistage purification process including ammonium sulfate fractionation, ion exchange chromatography on DEAE-sephacel, and gel chromatography on Sephadex G-200. The LDH preparation had a specific activity of 41.9 E/mg protein, with a purification rate of 101 and a yield of 26.8%. Its physical and chemical features were studied. The molecular weight of the native LDH molecule (148 kD) was determined, and the enzyme was shown to consist of four subunits, the molecular mass of which was determined by PAGE-Na electrophoresis in the presence of SDS-Na equal to 37 kD. The kinetic and regulatory properties, such as the Michaelis constants, the substrate inhibition constants, the effects of hydrogen-ion and temperature on the direct and reverse reactions, were studied.

Smilax moranensis M.Martens & Galeotti, Smilacaceae, root is a medicinal plant used among the Chatinos in Oaxaca, Mexico, to control type 2 diabetes. The objectives of the study were to isolate the bioactive compounds from the roots of Smilax and evaluate the chronic hypoglycemic effect of the ethanol–water extract. The main compounds were isolated from the methanolic extract via conventional phytochemical methods. The dried roots of S. moranensis were extracted with methanol and chromatographed on Sephadex LH 20. Fractions were chromatographed and purified on a silica gel chromatography column. The ethanol–water extract was orally administered to hyperglycemic rats for a period of 42 days, and glucose, glycated hemoglobin (HbA1c), and triacylglycerides were measured. Moreover, very-low-density lipoprotein was calculated. During the chemical investigation, three compounds were isolated and characterized, namely, 3-O-caffeoyl-quinic acid, 5-O-caffeoyl-quinic acid and trans-resveratrol, using various spectroscopic techniques. Animal experiments confirmed that the plant extract could control both the glucose and HbA1c levels. In conclusion, this study confirms that the roots of S. moranensis have hypoglycemic properties and suggests that the isolated compounds are potentially involved in this effect. Keywords: Type 2 diabetes, Hypoglycemic, Chlorogenic acids, Medicinal plants

Whey is recognized by the excellent nutritional and functional properties of its proteins, which can be hydrolyzed to peptides with an array of biological activities. In this work, sheep whey was treated with a novel proteinase from Bacillus sp. P7, resulting in hydrolysates with antioxidant and angiotensin-converting enzyme I (ACE-I) inhibitory activity. The hydrolysates were purified by gel filtration chromatography on Sephadex G-25 and the active fractions were subjected to ultrafiltration through 10 kDa cut-off membranes. The fraction enriched in bioactive peptides was encapsulated into phosphatidylcholine liposomes, which were characterized and evaluated by their ability to maintain biological activities. The physicochemical parameters of the liposomes showed constant values during 30 days, with mean diameter around 166 nm and zeta potential −17 mV. Both antioxidant and ACE-I-inhibitory activities were detected in the liposome preparations. In addition, the residual antioxidant activity was 87% after 30 days storage, suggesting the bioactive peptides are stable in this formulation. Liposome encapsulated peptides may offer the possibility of application in areas such as food science and technology, in order to increase the nutritional value and shelf life of food products, as well as for the development of functional foods.

A series of modified polysaccharide microparticles have been fabricated and their potential application for scavenging reactive oxygen species (ROS) and their derivatives to achieve osteoarthritis (OA) treatment has been explored. These microparticles were cross-linked dextran (Sephadex) with different carbazate substitution ratios determined by the TNBS assay and elemental analysis. It has been demonstrated that they could effectively scavenge carbonylated proteins and ROS including hydroxyl radicals (˙OH), superoxide anions (˙O2-) and H2O2 and their derivatives with high efficiency, improve the viability of H2O2-treated chondrocytes by reducing their ROS levels, as well as lower their inflammatory factors. The above ability of antioxidation and inflammation resistance improved with the increase of carbazate substitution ratio. Significantly, this work provided the proof that modified Sephadex successfully alleviated the deterioration of cartilage and the progression of OA in vivo. The proposed microparticles showed a very promising capability for reducing ROS levels and further treating OA.

Snakehead fish was a fish that has a high albumin content. Snakehead fish was used in the health sector known as medicinal freshwater fish, to accelerate the process of healing wounds after surgery and childbirth. The purpose of this study was to analyze the relative mobility (Rf) of the snakehead fish (Ophiocephalus striatus) albumin isolates by steaming at different temperatures and times. The research method for analyze the relative mobility (Rf) of the snakehead fish (Ophiocephalus striatus) albumin isolates by using gel filtration column chromatography, was detected by the SDS-PAGE electrophoresis method. Electrophoresis was carried out at a constant voltage of 125 v/ slab. Gel staining was carried out with Commasie Brilliant Blue R-250 in methanol-acetic acid-water. A standard protein mixture consisting of: myosin (200.0 kDa), β-galactosidase (116.3 kDa), phosphorylase B (97.4 kDa), bovine serum albumin (66, 2 kDa), ovalbumin (45.0 kDa), carbonic anhydrose (31.0 kDa), trypsin inhibitor (21.5 kDa), lysozyme (14.4 kDa) and aprotinim (6.5 kDa). The data analysis of this research was descriptive analysis to see the photos of the electrophoresis results. Meanwhile, for the analysis, the measurement of albumin isolation by gel filtration Sephadex G-75 was carried out with a divided plot design. The results showed that SDS-PAGE electrophoresis with the most complex amount of protein was albumin isolate, the effect of the steaming temperature was 40 oC for 30 minutes, located in 5 ml of the 1st fraction, 5 ml of the 2nd fraction and 5 ml of the 3rd fraction, with values of relative mobility (Rf) 0.0833 to 0.6944.

To study the bioactive potential of sulfated polysaccharide from green alga Ulva lactuca in Tuticorin coast. The isolated green alga was identified based on the morphological characters for the extraction of sulphated polysaccharide. The bioactive potential sulfated polysaccharide was determined using ABTS, free radical scavenging, antioxidant and its antibacterial activity was also evaluated. Sulfated polysaccharide (5.5 ± 0.6%) was extracted from green alga Ulva lactuca through the ethanol precipitation and purified through DEAE cellulose column and sephadex G -50. The superoxide anion and nitric oxide radical scavenging activity of sulfated polysaccharide was 71.74 ± 0.20% and 61.23 ± 0.48%, respectively. These results suggested that the in vitro antioxidant activity of sulfated polysaccharide obtained from Ulva lactuca can be employed for food industry. In addition to that, it possesses antibacterial activity against aquatic pathogens. This may also justify the frequent use of natural antioxidants in a variety of food products.

Despite the commercial availability of many different hydrogel formulations, the effective gel-based purification of single-walled carbon nanotubes (SWNT) remains exclusive to the gel Sephacryl S-200. In this study, 12 commercially available gels and two custom-synthesized gels were investigated for their ability to effectively purify SWNT, as determined through quantification of SWNT adsorption, elution, chiral selectivity, and overall process efficiency. The ability of each gel to separate SWNT was found to correlate with physiochemical properties, such as hydrogel pore size, the presence of ionic ligands, and both polysaccharide backbone and cross-linker compositions. While Sephacryl S-200 demonstrated superior separation efficiency and chiral selectivity among the gels studied, Superose 6 was found to adsorb more SWNT than Sephacryl S-200 per cm2 of the gel surface area and exhibited a unique preference for the (7,3) and (7,5) SWNT chiralities, in contrast to the established selectivity of Sephacryl S-200 for the (6,5) chirality. Collectively, this work both identifies gels that exhibit unique SWNT chiral selectivity and provides insights into the rational design of gels tailored for SWNT purification.

Bacteriocin has been identified as an excellent alternative to chemical preservatives due to its astonishing antimicrobial activity against food spoiling and food-borne pathogens. So there is a need to identify the newer and potent sources of bacteriocin producers. This study aims the isolation of potent bacteriocin producing microorganism from fresh fruits and vegetables, its production, purification, and characterization. Firstly, 43 isolates were analysed for its antimicrobial potential, out of which7 were found to inhibit the growth of various pathogens. Considering the results of antimicrobial activity; the microorganism isolated from mango was regarded as the most potent one; which was identified as Bacillus subtilis VS.70% ammonium sulphate precipitated and dialysed bacteriocin was purified using DEAE cellulose and sephadex G75 chromatography. Bacteriocin was purified by 24.64 fold with 8.65% recovery and its molecular weight was found to be 31.2kDa. The Purified bacteriocin was found to be stable at broad pH and temperature. It was found to be degraded by various proteases studied confirming its proteinaceous nature. Considering all these attributes; the purified bacteriocin isolated from Bacillus subtilis VS can be exploited by various food industries.

Research has shown that cartilage containing chondroitin sulfate and protein presents versatile bioactivities. Chondroitin sulfate in cartilage is beneficial to activate the immune system while the protein/peptide has not been fully understood. The current study investigated the antioxidant and anti-inflammatory properties of ethanol-soluble hydrolysates of sturgeon cartilage (ESCH) prepared through hot-pressure, enzymatic hydrolysis and ethanol extraction. UV spectrum, IR and agarose gel electrophoresis results suggested the successful exclusion of chondroitin sulfate from peptides. Nitric oxide (NO) floods in cells activated by inflammation. It was inhibited when administrated with ESCH. To further explain the observed anti-inflammatory activity, ESCH was separated with Sephadex G-15 into 3 components, among which F3 showed a higher NO inhibition rate and significantly reduced the production of the proinflammatory cytokine IL-6. In addition, the yield of IL-10 increased. Western blotting suggested that F3 downregulated the NO content and IL-6 level by suppressing Mitogen-activated protein kinases (MAPK) channels. Moreover, both ESCH and F3 showed DPPH and ABTS free radical scavenging abilities which was possibly related to the anti-inflammatory property. These results indicated that ESCH behaved anti-inflammatory and antioxidant activities. Cartilage may be a good source to produce anti-inflammatory peptides.

Repeated column chromatography using Sephadex LH-20, silica gel (SiO2), and octadecyl SiO2 (ODS) as well as preparative HPLC column chromatography led to isolation of a new dibenzocyclooctadiene lignan along with four known ones, gomisin L2 (1), L1 (2), M1 (3), and M2 (4). Their chemical structures were fixed based on MS, IR, and NMR data analyses. In addition, the stereochemistry of atropisomers, the absolute configuration of the axial chirality in a biphenyl structure, was confirmed by a CD experiment. The new lignan was named gomisin M3 (5).

Lycopi Herba has been broadly used as a traditional medicinal herb in Asia due to its ability to strengthen immunity. However, it is still obscure for its material basis and underlying mechanisms. Polysaccharide, as one of the most important components of most natural herbs, usually contributes to the immunomodulatory ability of herbs. Here, we aimed to detect polysaccharides from Lycopi Herba and examine their potential immunomodulatory activity. A novel polysaccharide (LHPW) was extracted from Lycopi Herba and purified by DEAE-52 cellulose chromatography and G-100 sephadex. According to physicochemical methods and monosaccharide composition analysis, LHPW was mainly composed of galactose, glucose, fructose, and arabinose. NMR and methylation analyses indicated that LHPW was a neutral polysaccharide with a backbone containing →3,6)-β-D-Galp-(1→, →4)-β-D-Galp-(1→ and →4)-α-D-Glcp-(1→, with the branches of →1)-β-D-Fruf-(2→ and →6)-α-D-Galp-(1→. Immunological tests indicated that LHPW could activate macrophage RAW264.7 and promote splenocyte proliferation. This study discovered a novel polysaccharide from Lycopi Herba and showed it was a potential immunomodulator.

In the present study, an extracellular esterase from Serratia sp. was purified 24.46 fold using an initial ammonium sulphate precipitation step (optimized concentration of 30–40%), followed by Diethylaminoethyl cellulose (DEAE-cellulose) chromatography and size exclusion Sephadex G-200 column chromatography steps. The molecular weight of the esterase using native polyacrylamide gel electrophoresis (PAGE) was determined to be 236 kDa and by using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) was found to be 60 kDa suggesting that the enzyme was a tetramer of 4 subunits. The purified esterase was able to catalyze the hydrolysis of p-nitrophenyl esters, especially p-nitrophenyl acetate. Maximum esterase activity was achieved in 0.15 M Tris–HCl buffer of pH 8.5 at 50 °C after 10 min. The enzyme was stable for at least 8 h at 4 and 35 °C but the half-life was determined to be 4.5 h at 50 °C and 3 h at 60 °C. The esterase activity was inhibited by detergents (1 mM) (Triton X-100, Tween 60, Tween 80, ethylenediamine tetraacetic acid and SDS) except Tween 20. The esterase activity was inhibited by organic solvents (1 mM) such as ethanol, methanol, acetone, acetonitrile and was stable in the presence of glycerol, isopropanol but the organic solvent dimethyl sulfoxide (DMSO) significantly (p