Partial Purification and Characterization of Bacteriocin-Like Inhibitory Substances Produced by Streptomyces sp. Isolated from the Gut of Chanos chanos.

Bacteriocin-like inhibitory substances (BLIS) have sparked great interest because of their promising use in food as natural antimicrobial agents. In this work, six Streptomyces isolates obtained from the gut of Chanos chanos demonstrated their ability to produce extracellular metabolites with inhibitory activity against Salmonella enterica serovar Typhimurium, Escherichia coli, Listeria monocytogenes, and Staphylococcus aureus. Exposure of the extracellular metabolites to proteolytic enzymes (i.e., proteinase-K, trypsin, and pepsin) revealed high sensitivity and confirmed their proteinaceous nature. The metabolites were stable at high temperatures (up to 100°C for 30 min) and a wide range of pH (pH 2.0-7.0). Fractionation of the crude BLIS by filtration yielded three fractions based on molecular weight: <3 kDa, 3-10 kDa, and >10 kDa. Analysis of the antibacterial activity of these fractions showed increased specific activity, especially in the fraction with a molecular weight (MW) of <3 kDa, relative to the crude sample. The fraction with MW < 3 kDa had minimum inhibitory and bactericidal concentrations in ranges 0.04-0.62 mg·mL-1 and 0.08-1.25 mg·mL-1, respectively. This fraction also showed better temperature and pH stability compared with crude BLIS. Brine shrimp toxicity assay revealed that this fraction has moderate toxicity with a 50% lethal concentration of 226.975 µg·mL-1 (i.e., moderate toxicity) to Artemia salina. Identification of the peptide sequences of this fraction by liquid chromatography-tandem mass spectrometry yielded 130 proteins with retention times of 15.21-19.57 min. Eleven proteins with MWs of 1345.66-2908.35 Da and composed of less than 30 amino acid residues with high hydrophobicity (15.34-26.22 kcal·mol-1) appeared to be responsible for the antibacterial activity of the fraction. This study revealed the potential application of BLIS from Streptomyces, especially BLIS SCA-8, as antibacterial agents.

Antifungal Activity and Major Bioactive Compounds of Water Extract of Pangium edule Seed against Aspergillus flavus.

Pangium edule seeds are widely used as spices in Southeast Asia in a fresh and fermented form and are reported to have active compounds for food preservation. However, scientific data on the active compounds of P. edule seed that can prevent the growth of toxigenic Aspergillus flavus have not been widely reported. This research subjected to determine the antifungal activity and identify the active compounds of water extract of old and fermented seed of P. edule against A. flavus. The water extract was compared to the extracts obtained by multilevel maceration using 50% ethanol, ethyl acetate, and n-hexane as solvents. Alkaloid, saponin, phenolic compound, flavonoid, triterpenoid, and glycoside were detected qualitatively in the crude extracts. The water extract showed the best activity to suppress the growth of A. flavus, determined by the agar dilution method, with the minimum inhibitory concentration (MIC) of 12.5 and 25 mg/mL for old and fermented seed, respectively. The water extracts showed a moderate toxicity with LC50 of 100-500 µg/mL, determined by the brine-shrimp toxicity test. After fractionation using 3 kDa molecular-weight (MW) cut-off ultrafiltration membrane, two fractions, i.e., fraction with MW < 3 kDa and >3 kDa, were obtained. The fraction with MW < 3 kDa showed the best antifungal activity with the MIC of 6.25 and 12.5 mg/mL for old and fermented seed, respectively. LC-MS/MS profile showed that different compounds belong to fatty acid, amino acid, glycoside, and peptide were found as major active compounds in the fractionated water extract. The principal compounds and partial least-square analysis, however, suggested that fatty acid and glycoside are responsible for the antifungal activity. Hence, this study concluded that the water extract of P. edule seed had promising antifungal activity against A. flavus which was due to presence of particular compounds belong to fatty acid and glycoside.

Characterization of Streptomyces Isolates Associated with Estuarine Fish Chanos chanos and Profiling of Their Antibacterial Metabolites-Crude-Extract.

Streptomyces has been reported as an essential producer of bioactive substances, including antibiotics and other types of antimicrobials. This study investigated antibacterial-producing Streptomyces isolated from the gut of estuarine fish Chanos chanos, emphasizing screening for the producer of peptide-containing antibacterial compounds. Eighteen isolates were found during preliminary screening, in which four isolates showed the best antibacterial activities. Based on the morphological, physiological, and biochemical characterization, as well as 16S rRNA partial sequencing, all of the four isolates belonged to Streptomyces. Three isolates were suspected as novel isolate candidates based on homology presentations and phylogenetic tree analysis. Disk-diffusion assay of the metabolite-crude-extract from the isolates showed broad-spectrum inhibitory activities against Staphylococcus aureus ATCC 25923, Bacillus cereus ATCC 10876, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa InaCC B52 with minimum inhibitory concentration and minimum bactericidal concentration ranging from 2.5-10 mg/mL and 5-10 mg/mL, respectively. The highest antibacterial activity with low MIC and MBC values was shown by isolate AIA-10. Qualitative HPLC profiling revealed that the metabolic-crude-extracts showed many peaks with intensive area at 210 and 214 nm, especially from SCA-11 and AIA-10, indicating the presence of peptide groups in the structure of the constituent compound. The results also suggested that crude extracts SCA-11 and AIA-10 had higher hydrophobicity properties than the other extracts. Further characterization of the active compound was needed to find out which compounds were responsible for the antibacterial activity. The results of this study indicated that some Streptomyces isolated from new environmental niches, i.e., gut of estuarine fish Chanos chanos, produce promising peptide-containing bioactive compounds.

Identification and Characterization of a-Glucosidase Inhibition Flavonol Glycosides from Jack Bean (Canavalia ensiformis (L.) DC.

Although the intake of jack bean (Canavalia ensiformis (L.) DC.), an underutilized tropical legume, can potentially decrease the risk of several chronic diseases, not much effort has been directed at profiling the polyphenolics contained therein. Hence, this work aimed to identify and quantify the dominant jack bean polyphenolics, which are believed to have antioxidant and other bioactivities. Four major compounds were detected and identified as kaempferol glycosides with three or four glycoside units. Their structures were established based on UV-visible, 1d, 2D NMR, and HR-ESI-MS analyses. Specifically, kaempferol 3-O-a-l-rhamnopyranosyl (1®6)- b-d-glucopyranosyl (1®2)-b-d-galactopyranosyl-7-O-[3-O-o-anisoyl]-a-l-rhamnopyranoside was detected for the first time, while the other three compounds have already been described in plants other than jack bean. This new compound was found to have a higher a-glucosidase inhibition activity compared to acarbose.

Low molecular weight compounds responsible for savory taste of Indonesian soy sauce.

Indonesian soy sauce is made using only soybeans as the nitrogenous source. Moromi obtained from fermentation of yellow soybeans using Aspergillus sojae as the starter was investigated. The fraction with molecular weights of less than 500 Da obtained by stepwise ultrafiltration was then fractionated by several chromatographic procedures, including gel filtration chromatography and RP-HPLC. Several chemical analyses, CE profiles, and taste profiles were performed to obtain the most intense umami fraction. The main components eliciting or enhancing the umami taste present in the fraction were purified and identified by protein sequencing, ESI-MS, and (1)H NMR at 400 MHz. Besides free l-glutamic acid and aspartic acid, free aromatic amino acids such as l-phenylalanine and l-tyrosine may also play an important role in impressing savory or umami taste of Indonesian soy sauce at their subthreshold concentrations and in the presence of salt and free acidic amino acids. This is reported as a new phenomenon of the so-called bitter amino acids.