A Novel Agarase, Gaa16B, Isolated from the Marine Bacterium Gilvimarinus agarilyticus JEA5, and the Moisturizing Effect of Its Partial Hydrolysis Products

We recently identified a β-agarase, Gaa16B, in the marine bacterium Gilvimarinus agarilyticus JEA5. Gaa16B, belonging to the glycoside hydrolase 16 family of β-agarases, shows less than 70.9% amino acid similarity with previously characterized agarases. Recombinant Gaa16B lacking the carbohydrate-binding region (rGaa16Bc) was overexpressed in Escherichia coli and purified. Activity assays revealed the optimal temperature and pH of rGaa16Bc to be 55 ∘C and pH 6–7, respectively, and the protein was highly stable at 55 ∘C for 90 min. Additionally, rGaa16Bc activity was strongly enhanced (2.3-fold) in the presence of 2.5 mM MnCl2. The Km and Vmax of rGaa16Bc for agarose were 6.4 mg/mL and 953 U/mg, respectively. Thin-layer chromatography analysis revealed that rGaa16Bc can hydrolyze agarose into neoagarotetraose and neoagarobiose. Partial hydrolysis products (PHPs) of rGaa16Bc had an average molecular weight of 88–102 kDa and exhibited > 60% hyaluronidase inhibition activity at a concentration of 1 mg/mL, whereas the completely hydrolyzed product (CHP) showed no hyaluronidase at the same concentration. The biochemical properties of Gaa16B suggest that it could be useful for producing functional neoagaro-oligosaccharides. Additionally, the PHP of rGaa16Bc may be useful in promoting its utilization, which is limited due to the gel strength of agar.

Prebiotics Inulin Metabolism by Lactic Acid Bacteria From Young Rabbits

Inulin as a commercial prebiotic could selectively promote the growth of beneficial gut microbes such as lactic acid bacteria (LAB). Whether LAB in rabbit gut possesses the capability to metabolize and utilize inulin is little known. Therefore, this study recovered 94 LAB strains from neonate rabbits and found that only 29% (28/94) could metabolize inulin with both species- and strain-specificity. The most vigorous inulin-degrading strain, Lacticaseibacillus paracasei YT170, could efficiently utilize both short-chain and long-chain components through thin-layer chromatography analysis. From genomic analysis, a predicted fosRABCDXE operon encoding putative cell wall-anchored fructan β-fructosidase, five fructose-transporting proteins and a pts1BCA operon encoding putative β-fructofuranosidase and sucrose-specific IIBCA components were linked to long-chain and short-chain inulin utilization respectively. This study provides a mechanistic rationale for effect of inulin administration on rabbits and lays a foundation for synbiotic applications aimed at modulating the intestinal microbiota of young rabbits.

Chemical analysis of callus extracts from toxic and non-toxic varieties of Jatropha curcas L.

Jatropha curcas L. belongs to Euphorbiaceae family, and it synthesizes flavonoid and diterpene compounds that have showed antioxidant, anti-inflammatory, anticancer, antiviral, antimicrobial, antifungal and insecticide activity. Seeds of this plant accumulate phorbol esters, which are tigliane type diterpenes, reported as toxic and, depending on its concentration, toxic and non-toxic varieties has been identified. The aim of this work was to characterize the chemical profile of the extracts from seeds, leaves and callus of both varieties (toxic and non-toxic) of Jatropha curcas, to verify the presence of important compounds in dedifferentiated cells and consider the possibility of using these cultures for the massive production of metabolites. Callus induction was obtained using NAA (1.5 mg L−1) and BAP (1.5 mg L−1) after 21 d for both varieties. Thin layer chromatography analysis showed differences in compounds accumulation in callus from non-toxic variety throughout the time of culture, diterpenes showed an increase along the time, in contrast with flavonoids which decreased. Based on the results obtained through microQTOF-QII spectrometer it is suggested a higher accumulation of phorbol esters, derived from 12-deoxy-16-hydroxy-phorbol (m/z 365 [M+H]+), in callus of 38 d than those of 14 d culture, from both varieties. Unlike flavonoids accumulation, the MS chromatograms analysis allowed to suggest lower accumulation of flavonoids as the culture time progresses, in callus from both varieties. The presence of six glycosylated flavonoids is also suggested in leaf and callus extracts derived from both varieties (toxic and non-toxic), including: apigenin 6-C-α-L-arabinopyranosyl-8-C-β-D-xylopyranoside (m/z 535 [M+H]+), apigenin 4′-O-rhamnoside (m/z 417 [M+H]+), vitexin (m/z 433 [M+H]+), vitexin 4′-O-glucoside-2″-O-rhamnoside (m/z 741 [M+H]+), vicenin-2 (m/z 595 [M+H]+), and vicenin-2,6″-O-glucoside (m/z 757 [M+H]+).

Pemisahan Senyawa Aktif Fraksi Petroleum Eter dan Etil Asetat Hasil Hidrolisis Ekstrak Etanol Hydrilla verticillata dari Ranu Grati Pasuruan

<p><em><span lang="EN">Hydrilla verticillata</span></em><span lang="EN"> is one of water plants that has some bioactivities. The </span>presence of secondary metabolites in <em>H. verticillata</em> is responsible for the bioactivity. The purpose of this study was to determine and separate bioactive compounds from fraction of petroleum ether (PE) and ethyl acetate (EA) as a result of hydrolysis of <em>H. verticillata</em> ethanol extract. <em>H. verticillata</em> was extracted by maceration method using ethanol solvent, hydrolyzed with hydrochloric acid and partitioned respectively with petroleum ether and ethyl acetate. Crude ethanol extract, PE and EA fraction were identified their secondary metabolites. The phytochemical test results showed <em>H. verticillata</em> ethanol extract containing alkaloid compounds, flavonoids, tannins, saponins, triterpenoids, and steroids. PE fraction contained steroids and terpenoids, while EA fraction contained flavonoids, steroids, and triterpenoids. Analytical thin layer chromatography analysis showed n-hexane : ethyl acetate (4: 1) eluent as the best mobile phase for separating steroids. The preparative thin layer chromatography analysis of <em>H. verticillata</em> fraction using n-hexane : ethyl acetate (8: 2) as mobile phase resulted in 17 and 14 spots of PE and EA fractions, respectively.</p><p>Keywords: <em>Hydrilla verticillata</em>, thin layer chromatography, phytochemicals<em></em></p><p> </p><p><em>Hydrilla verticillata</em> merupakan salah satu tanaman air yang banyak memiliki bioaktivitas. Adanya metabolit sekunder pada H. verticillata yang bertanggung jawab terhadap bioaktivitas ini. Tujuan penelitian ini adalah untuk mengetahui <span lang="IN">dan memisahkan </span>senyawa aktif dari <span lang="IN">fraksi petroleum eter</span> (PE) <span lang="IN">dan etil asetat</span> (EA) <span lang="IN">hasil hidrolisis </span>ekstrak <span lang="IN">etanol </span>H. verticillata. Ekstrak diperoleh dengan metode maserasi menggunakan pelarut etanol, dihidrolisis dengan asam klorida dan dipartisi masing-masing dengan petroleum eter dan etil asetat. Ekstrak kasar etanol, fraksi PE dan EA diuji kandungan metabolit sekundernya. Hasil <span lang="IN">u</span>ji fitokimia <span lang="IN">menunjukkan </span>ekstrak<span lang="IN"> etanol</span>H. verticillata mengandung senyawa alkaloid, flavonoid, tanin, saponin, triterpenoid, dan steroid. Fraksi PE mengandung steroid dan terpenoid, sedangkan fraksi EA mengandung flavonoid, steroid, dan triterpenoid. Hasil analisis KLTA menunjukkan eluen n-heksana : etil asetat (4:1) sebagai fasa gerak terbaik untuk memisahkan steroid.Hasil analisis KLTP fraksi H. verticillatamenggunakan perbandingan fasa gerak n-heksana : etil asetat (8:2) menghasilkan spot fraksi PE dan EA berturut-turut sebanyak 17 dan 14 spot.</p><p><span>Kata Kunci: </span><em><span>Hydrilla verticillata</span></em><span>, kromatografi lapis tipis, uji </span><span lang="IN">fitokimia</span></p>