Vacuum Liquid Chromatography Research Articles

Antimicrobial and Antioxidant Activities of Novel Marine Bacteria (Bacillus 2011SOCCUF3) Isolated from Marine Sponge (Spongia officinalis)

Background : Bacillus species represent a rich source of new bioactive metabolites that can combat diseases. Methods: Bacillus strain was isolated from the marine sponge Spongia officinalis and routinely maintained on marine broth. The bacteria strain was identified as Bacillus 2011SOCCUF3 using 16S rDNA sequencing. The strain was cultured on Tryptone Casein Oat Soluble Starch (TCOATSS) media with continuous agitation for 4 days. The fermented broth was centrifuged, and the supernatant was mixed with 10% (w/v) of adsorbent resin (XAD-7HP and XAD-16N, 1:1) and shaken continuously at a reduced speed for 7 h; and the resin was collected by filtration through sintered glass funnel and washed with MilliQ water, and then eluted with methanol to obtain the extract. The extract was evaporated in vacuo at reduced temperature and pressure to obtain the dry extract. The dry extract was purified by vacuum liquid chromatography, eluting with methanol in acetone gradient. The in vitro antimicrobial and antioxidant activities were investigated using the agar-well diffusion, DPPH scavenging and the phosphomolybdate methods respectively. Results: The extract and fractions showed good antimicrobial activities with minimum inhibitory concentration range of <1.0 mg/mL. The extract and fractions also exhibited good antioxidant activities with their IC50 values been comparable to the standard. Conclusion: Thus, a novel Bacillus strain isolated from the marine sponge (Spongia officinalis) obtained from Cortiou and Riou, France, exhibited promising antimicrobial and antioxidant activities.

Physicochemical and antimicrobial assessment of Iranian Propolis gathered in Qazvin province

Background: Propolis is one of the useful bee colony products that have been used in traditional medicine for centuries. In this study, the physicochemical characters and their antibacterial effect of Iranian Propolis collected from Qazvin province was assessed.Methods: In this study, Thin Layer Chromatography and Vacuum Liquid Chromatography to detect different compounds of the extract have been used. In the initial evaluation of Propolis extract, it was found that the extract includes variable compounds with different polarity; so, the initial classification of extract with different polarity solvents was essential. Finally, 0.1 gr hydro alcoholic Propolis was injected to the HPLC by ultrasound. The antibacterial effect of Iranian ethanol extract Propolis was measured using a microdilution method against Pseudomonas aeruginosa: P. aeruginosa and Staphylococcus aureus: S.aureus standard strains and the minimum bactericidal and inhibitory concentration were defined.Results: Primary analysis of the ethanol extract by analytical Thin Layer Chromatography, demonstrated the presence of flavonoid and phenol in it. Minimum inhibitory concentration and Minimum Bactericidal Concentration for Staphylococcus aureus: S.aureus standard strain was 2.5mg/ml. The same procedure was done for Pseudomonas aeruginosa: P. aeruginosa standard strain and the Minimum inhibitory concentration and Minimum Bactericidal Concentration were 50mg/ml of Propolis extracts. Conclusion: According to the results, the alcoholic extract of propolis from Qazvin province of Iran provides significant antimicrobial activity. Its powerful activity may be due to high total phenolic and flavonoid contents.Keywords: Iranian propolis, Antibacterial activity, Phenolic compounds, Flavonoid compound

Antibacterial Activity of Scopoletin from Stem Bark of Aleurites moluccana Against Salmonella typhi

Background: Based on a previous survey of ethnopharmaceutical studies (local culture), it was known that the use of decoction of candlenut stem bark (Aleurites Moluccana) was used as typhoid medicine in the treatment tradition of the people of East Kalimantan. In antibacterial screening by diffusion test, methanol extract of stem bark A.moluccana showed excellent growth inhibition of Salmonella thypimurium. Working method: Simplicia of A.moluccana stem bark was extracted by maceration using methanol. Maserate was extracted liquid-liquid using n-hexane and ethyl acetate solvents. The hexane and ethyl acetate fraction was monitored for active spots with TLC bioautography. The active compound is separated using vacuum liquid chromatography and radial chromatography. Results: The fraction J of KCV and J2 subfraction results from radial chromatography provides antibacterial activity of S. thpimurium. From the J2 subfraction obtained pure isolates in the form of yellowish needle crystals. The isolate was tested for antibacterial S.thypimurium using the microdilution method with a value of MIC is 250 µg/ml. Based on spectroscopic data and comparing the published spectra of the compound, the elucidation of the isolate is Scopoletin (7-hydroxy-6-methoxycoumarin).

A bioactive compound isolated from Duku ( Lansium domesticum Corr) fruit peels exhibits cytotoxicity against T47D cell line.

Background: Breast cancer is a major health problem for women globally. Many attempts have been promoted to cure cancer by finding new anticancer medicines from natural resources. Despite the richness of biodiversity discovered, there are some natural resources that remain unexplored. Fruit peels of Duku ( Lansium domesticum Corr.) are rich with compounds that may have the potential to be developed as anticancer drugs. This study aimed to isolate cytotoxic compounds from the fruit peels of L. domesticum and assess their cytotoxic nature against T47D cells. Methods: Powdered peels were macerated with ethyl acetate and the filtrate was evaporated to give EtOAc extract A. Dried extract A was triturated with n-hexane to give n-hexane soluble fraction B and insoluble fraction C. The cytotoxic nature of these three samples were assessed using MTT assay using T47D cells and doxorubicin as a control. Results: Fraction C that showed the smallest IC50 (25.56 ± 0.64μg/mL) value compared to extract A and fraction B. Fraction C was further fractionated by vacuum liquid chromatography to give 6 subfractions. Subfraction 2 showed a single compound based on thin layer chromatography, and this compound was identified as Lamesticumin A on the basis of its spectroscopic data. Lamesticumin A demonstrated cytotoxic activity against T47D cell lines with an IC50 value of 15.68± 0.30µg/mL. Conclusions: Further research is needed to investigate the potential of the natural compound Lamesticumin A derived from L. domesticum fruit peel as an anticancer therapy.

Anti-Adipogenic compound from Guazuma ulmifolia Leaf

The purpose of this research was to test the anti-adipogenic activity and to identify the active compound of the leaf of Guazuma ulmifolia. The activity of anti-adipogenic of the compound of G. ulmifolia leaf was tested using primary cultures of Wistar strain white mouse preadipocytes. Isolation of the active compound was carried out in the following way. The powder of G. ulmifolia leaf was defated with n-hexane followed by extraction with 70% ethanol. The extract was concentrated and fractionated by liquid-liquid partition using chloroform and ethyl acetate solvents. The ethyl acetate fraction was loaded into a Vacuum Liquid Chromatography (VLC) followed by a reverse phase preparative layer chromatography to obtain a pure isolate. Isolated compound inhibited preadipocytes proliferation and differentiation. Intensive spectroscopic and spectrometric analysis suggested molecular structure of a tiliroside which agreed with previously reported data. The study successfully revealed the traditional claims of the anti-obesity agent of a leaf of G. ulmifolia through inhibition of preadipocytes proliferation and differentiation.

Bioactivity-directed isolation of antisickling compounds from Cnidoscolus acontifolius (Mill.) I.M. Johnst leaf extract

Context: There is continuous search for therapeutic agents from indigenous plants that can be employed in the treatment of sickle cell anemia. Aims: To evaluate the antisickling potential of Cnidoscolus aconitifolius leaf extract, determine the most active fraction, and isolate the putative compounds. Methods: Oven dried leaves of C. aconitifolius (CA) were extracted by maceration in ethanol for 72 h. The extract was fractionated into n-hexane, dichloromethane, ethyl acetate and methanol using vacuum liquid chromatography (VLC). The crude CA extract and fractions were subjected to inhibitory and reversal antisickling assays at 0.25-4.00 mg/mL concentration range. Bioactivity-directed fractionation of the most active fraction was done on repeated silica gel column chromatography, followed by preparative thin-layer chromatography, and their analyses on thin-layer chromatography. The isolated compounds were characterized using spectroscopic methods of 1H- and 13C- Nuclear Magnetic Resonance, COSY, HMBC, HSQC, and LC-MS. Results: The results showed that CA had 80.4 ± 0.15% inhibitory and 56.0 ± 2.90% reversal effects at 4 mg/mL. The ethyl acetate fraction gave significantly higher (p<0.05) inhibitory (68.0 ± 4.32%) and reversal (61.4 ± 6.2%) activities at 4 mg/mL than the other VLC fractions. The positive control Ciklavit® had 59.8 ± 0.3% inhibitory and 56.6 ± 0.2% reversal properties. Two compounds, T1 and T2 were isolated from the ethyl acetate fraction and identified as tetramethyl bicosahydropicen-3-ol and 5β-pregnane, respectively. Compound T1 demonstrated an inhibitory effect of 83.6 ± 0.11%. Conclusions: The study concluded that the ethyl acetate fraction of the ethanol extract of C. aconitifolius has the highest antisickling property and identified tetramethylbicosahydropicen-3-ol as a potential antisickling agent.

Isolation of Two Isochlorogenic Acid Isomers from Phenolic Rich Fraction of Artemisia turanica Krasch.

Total phenolic content (TPC) and antioxidant capacity of five different extracts (petroleum ether (40-60), dichloromethane, ethyl acetate, ethanol and ethanol-water (1:1 v/v)) of Artemisia turanica (A. turanica) aerial parts were determined and phytochemical study on the most promising extract was carried out. Folin–Ciocalteu method, 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging test, β-carotene bleaching (BCB) method, and ferrous ion chelating (FIC) assay were performed. Vacuum liquid chromatography (VLC) and semi-preparative HPLC were used for bioassay-guided phytochemical isolation. Structures of isolated compounds were established using spectroscopic analysis including NMR and MS. Among all the extracts analyzed, the hydroethanolic extract exhibited the highest phenolic content and antioxidant activity. VLC of this extract yielded seven fractions (A to G) which were subjected to all antecedent experiments. The same sample (Fraction D) showed the highest total phenolic content and free radical scavenging activity but the only statistically significant correlation between TPC and EC50 values was observed for BCB. 3,5-dicaffeoylquinic acid (isochlorogenic acid A), and 4,5-dicaffeoylquinic acid (isochlorogenic acid C) was isolated from the most active fraction. Antioxidant activity of A. turanica is probably partly due to the presence of isomers of isochlorogenic acid.

Antitumor activity of methoxylated flavonoids separated from Achillea fragrantissima extract in Ehrlich’s ascites carcinoma model in mice

Introduction: Achillea fragrantissima extract has been used in folk medicine as an anticancer. The present study describes the antitumor activities of the extract and the flavonoid compounds and attempts to explain the mechanisms underlying these activities. Methods: The whole plant of A. fragrantissima was collected and ground to produce a fine powder which subjected to the maceration process. Step gradient elution was used using silica gel vacuum liquid chromatography (VLC). The fractions were compared by thin layer chromatography (TLC) and concentrated. Final purification was performed using Sephadex LH-20 to give three compounds cirsiliol, chrysosplenol D and cirsimaritin from F4 and one compound, eupatilin-7-methyl ether from F6. In this study, Ehrlich’s ascites carcinoma (EAC) model was used as the model of cancer. Results: Achillea fragrantissima extract and its isolated methoxylated flavonoids significantly reduced the weight of tumor discs compared to EAC-control group. In addition, cirsimaritin and eupatilin 7-methyl ether treatments produced a dose-dependent reduction in tumor weight. Serum tumor necrosis factor-alpha (TNF-ɑ) level showed that A. fragrantissima extract and its isolated methoxylated flavonoid compounds significantly reduced its serum level compared to the EAC-control group. Furthermore, A. fragrantissima extract and the flavonoids significantly increased the Total antioxidant capacity (TAC) compared to EAC-control. A. fragrantissima extract and its isolated methoxylated flavonoids produced a better differentiation of tumor cells, with reduced nuclear pleomorphism and better formed tubular structures especially with high dose indicating the induction of apoptotic mechanism. Conclusion: Achillea fragrantissima extract and its isolated methoxylated flavonoids exhibit antitumor activities that may be attributed to the antioxidant properties and the induction of the apoptotic process.

Antibacterial Activity of Benzyl Benzoate and Crotepoxide from <i>Kaempferia rotunda</i> L. Rhizome

Benzyl benzoate and crotepoxide are the major components of Kaempferia rotunda L. rhizome. However, the bioactivity study of benzyl benzoate and crotepoxide as the antibacterial activity were still limited. Therefore, the antibacterial activity of benzyl benzoate and crotepoxide against four pathogenic bacteria, i.e., Escherichia coli ATCC 25922, Enterococcus aerogenes ATCC 13048, Bacillus cereus ATCC 6538 and Staphylococcus aureus ATCC 11778 were investigated. The isolation steps were include the extraction by maceration with acetone, then the acetone extract was partitioned with n-hexane:methanol (1:1) and ethyl acetate:water (1:1) respectively. The isolation by liquid vacuum chromatography followed by column chromatography was obtained benzyl benzoate from the n-hexane fraction and crotepoxide from ethyl acetate fraction. The molecular structure of isolated compounds was identified based on NMR (1D and 2D) spectroscopic data. The antibacterial activity assay of isolated compounds was carried out using the disc diffusion method. The antibacterial evaluation confirms that the benzyl benzoate and crotepoxide exhibits a medium level activity. Benzyl benzoate showed highest antibacterial activity against B. cereus at the MIC of 50 μg/mL and inhibitory zone of 5.9 mm, while the crotepoxide showed highest antibacterial activity against E. aerogenes at the MIC of 100 μg/mL with inhibitory zone 6.1 mm.

Isolation and Identification of DPPH Radical (2,2-diphenyl-1-pikrylhidrazyl) Scavenging Active Compound in Ethyl acetat fraction of Piper acre Blume

Piper acre Blume, known as Black Betel (local name), is a plant that is widely used by the people of East Kalimantan, especially in Samarinda, for the treatment of illness. Leaves (3-4 months old) are collected from Samarinda, extracted, fractionated, and monitored by DPPH antiradical activity. The isolation of the Piper acre Blume is performed on the active fraction, and the structure identification is based on spectroscopic data of the compound. The leaves were dried, pulverized, and macerated with MeOH. Dried MeOH extract was obtained upon evaporation of the solvent. The extract was then fractionated by vacuum liquid chromatography (vlc), eluted gradually by solvents having different polarities (n-hexane, ethyl acetate and methanol). The fractions obtained were monitored using TLC [n-hexane: ethyl acetate (3: 1 v/v)] that was visualized by UV254 nm, UV366 nm and DPPH. The isolation was performed by preparative TLC [SiO2, n-hexane: ethyl acetate (3: 1)] on ethyl acetate fraction that showed the highest DPPH antiradical value. A single compound was obtained, and it appeared as a round spot and pure according to TLC performances at 3 different solvent systems. The isolated Piper acre Blume compound displayed the IC50 value on the anti-radical DPPH (measured at λ 520 nm) as 10.41µg/mL. The IR spectrum (KBr) showed –OH band (3450 cm-1), aliphatic bands [alkene, 3010 cm-1; alkana 2900 cm-1), an aromatic overtone bands (1900-200 cm-1) and a strong C=O band (1725 cm-1). The NMR (1H- and 13C-) (mono and 2D) indicated the present of a p-di-substituted aromatic signals (δ, 7.54 and 7.52, d, J =6 Hz, 1 H each), 2 methyl (δ, 0.96, d, J = 7.0 Hz, 6 Hs), a triplet signal (δ, 4.22 ppm). Other signals of CH- and CH2 were shown as m signals at δ, 1.64 and 1.34 ppm. Based on those data, the compound was identified as isoamyl p-OH benzoate that is grouped as parabens used as a preservative in the pharmaceutical preparations. In conclusion, the anti-radical (DPPH) active compound present in the leaves of Piper acre Blume is identified as isoamyl p-OH benzoate, having IC50 value anti-radical DPPH 10,41µg/mL.

Enhanced process integration for the extraction, concentration and purification of di-acylated cyanidin from red cabbage

Anthocyanins are currently one of the most important pigments in the plants with great proven health-promoting benefits, however, their unstable chemical nature makes difficult their purification process. This work proposes the integration of two stages for the separation and purification of the pigments from red cabbage leaves. For the first stage, the acidified extract, containing 32.05 mg cyanidin-3-glucoside per milliliter (mg ECyn-3-glu•L−1), was subjected to micro, ultrafiltration and adsorption processes, allowing to concentrate up to 3221.45 mg ECyn-3-glu•L−1 of the pigments after an adsorption process. In the second stage, the pigments were fully fractionated by molecular exclusion chromatography, reverse-phase vacuum liquid chromatography and semi-preparative chromatography, isolating the stable di-acylated cyanidin with antioxidant activity. Finally, purified fractions were also analyzed by NMR analysis.

Anti-Oxidant, Anti-Malarial, and Phytochemical Studies on Muscari inconstrictum Bulbs Distributed in Iran

Background: Muscari inconstrictum Rech. f. is an ornamental and bulbous species with various medicinal and biological effects. Objectives: This study was designed to evaluate the anti-oxidant and anti-malarial activities of M. inconstrictum as one of the Iranian species of Muscari genus. Additionally, preliminary phytochemical investigation of the extracts with different polarity was performed. Methods: M. inconstrictum bulbs were extracted with n-hexane, chloroform, and methanol (MeOH) by Soxhlet apparatus, in the order of their polarity. Next, vacuum liquid chromatography (VLC) was used for the fractionation of extracts. Free radical scavenging and anti-malarial activities were investigated via DPPH (2,2-diphenyl-1-picrylhydrazyl) and cell-free β-hematin formation methods. Thin layer chromatography (TLC) and gas chromatography-mass spectrometry (GC-MS) were used for the characterization of potent fractions. Results: Among different extracts of M. inconstrictum, bulbs, chloroform, and n-hexane extracts were the most potent anti-oxidant and anti-malarial extracts, respectively. Moreover, methanolic, 80% and 100% ethyl acetate/n-hexane VLC fractions of chloroform extract showed significant anti-oxidant activities, which can be related to the presence of flavonoid and coumarin structures. Furthermore, 40% ethyl acetate/n-hexane VLC fraction of n-hexane extract with saponin structures is introduced as the most potent anti-malarial part. GC-MS analysis of methanol fraction of chloroform extract, 40% ethyl acetate/n-hexane VLC fraction of n-hexane extract, and the volatile oil of plant demonstrated the presence of phenolic monoterpenoid, fatty acid derivatives, and sesquiterpenoid structures as the main ingredients, respectively. Conclusions: It seems that more studies on M. inconstrictum are necessary to focus on pure compounds and their biological activities.

Isolation of Four Flavanones from the Leaves of Macaranga Hypoleuca (Rchb.f. & Zoll.) Müll.Arg

Macaranga is a large genus belongs to Euphorbiaceae family which commonly distributed in the tropical region of Africa, South-East Asia, China, and India. The plant of this genus contain flavonoids and stilbenes bearing various terpenyl groups including prenyl, geranyl and farnesyl groups. The leaves of M. hypoleuca were collected from reserved forest UiTM Jengka Pahang, Malaysia, and the dried powdered leaves were macerated in methanol at room temperature. The crude methanol extract was subjected to liquid-liquid partition using n-hexane and ethyl acetate to obtain hexane, ethyl acetate and aqueous fractions. The ethyl acetate fraction was semi purified using vacuum liquid chromatography (VLC) to give nine major fractions (MHL1-MHL9). Fraction MHL3 was further purified by column chromatography (CC) and preparative thin layer chromatography (p-TLC) to give two pure compounds, 8-prenylnaringenin (1) and sakuranetin (5,4’-dihydroxy-7-methoxyflavanone) (3). Meanwhile, fraction MHL6 was purified using column chromatography (CC) and p-TLC to yield another two pure compounds, 6-(3-hydroxy-3-methyl)naringenin (2) and 7-O-methyleriodictyol (4). The chemical structure of these isolated compounds were determined based on their 1D and 2D NMR, UV, and IR data. From this study, four flavanones were isolated from the leaves of M. hypoleuca.

Revelation of New Compound from Ethanolic Extract of Fragaria x ananassa var. Lembang

Fragaria x ananassa (strawberry) is a subtropical plant that can adapt well in tropical highlands. Fragaria x ananassa have been widely used to cope with health problems. The active compound component of secondary metabolites contained in Fragaria x ananassa has the potential as an antioxidant. This research is done to isolate secondary metabolites from extract of Fragaria x ananassa fruits. Extract Fragaria x ananassa was produced by maceration using ethanol as the solvent. Separation and isolation compound were carried out using Vacuum Liquid Chromatography (VLC) and Gravity Column Chromatography (GCC) guided by Thin Layer Chromatography (TLC) using hexane: ethyl acetate (3:7) as the eluent. The flavonoid compound was determined by the total content of phenolic and flavonoid in extract of Fragaria x ananassa fruits. The results of total phenolic content and total flavonoid content were 0.1130 mg/g and 0.0112 mg/g, respectively. The alkaloid compound was determined by Dragendorff testing. The elucidation of the structure by Fourier Tansform Infrared (FTIR), Nuclear Magnetic Resonance (NMR), and Liquid Chromatography Mass Spectrometry (LCMS) showed that the active compound contained in the secondary metabolite of extract ethanol from Fragaria x ananassa is 3-Cyclopentyl-5-(1-hydroxyethyl)-1,6-dihydro-7H-pyrazolo[4,3- d]pyrimidin-7-one.Keywords: Fragaria x ananassa extract, flavonoid, alkaloid, total phenolic and flavonoid content, FTIR, NMR, LCMS.

GC-MS Analysis and Attenuation of Bleomycin-Induced Lung Fibrosis by Fractionated Methanolic Extract of Cissampelos Owariensis in Wistar Rats

ABSTRACTCrude methanolic leaf extract of Cissampelos owariensis was fractionated using vacuum liquid chromatography and analyzed by gas chromatography–mass spectrometry (GC-MS). Male Wistar rats were induced pulmonary fibrosis (PF) by a single intraperitoneal administration of 0.75 U 100 g−1 bleomycin (BLM), divided equally into four groups and given orally fresh water (control), 10 mg kg−1 quercetin, 200, or 400 mg kg−1 crude methanol leaf extract (crudemet) of C. owariensis daily for 28 d. Plasmatic enzymes, antioxidants, and lipid peroxidation (LPO) were determined spectrophotometrically, and the histology of lung section was done microscopically. GC-MS revealed the presence of hexadecanoic, phytol, 9,12-octadecadienoic acid (z,z)-methyl ester. Lung fibrosis increased the plasmatic total protein and alkaline phosphatase (ALP) activities. The BLM-administered rats showed decreased sodium dismutase (SOD), glutathione (GSH) activity, and alveolar hyperplasia. Oral administration of crudemet of C. owariensis ameliorated total protein, ALP activity, and LPO levels. The bioactive agents present in the C. owariensis extract may have the potential of reducing pathophysiology through increased antioxidants, thereby reducing BLM-induced lung hyperplasia.

Gartanin Compounds from Extract Ethanol Pericarp Mangosteen (Garcinia mangostana Linn.).

AIM:The study aimed to isolate and identification secondary metabolite from pericarp Garcinia mangostana Linn.METHODS:The first step of this research was maceration of sample using alcohol 70% solvent. The separation and purification of compounds using Vacuum Liquid Chromatography (VLC), Radial Chromatography (RC). The purity of isolate was analyzed by thin layer chromatography (TLC) and melting point. Compounds identified using spectroscopi IR, NMR-1D (1H, 13C-NMR and DEPT) and 2-D NMR (HMQC and HMBC).RESULTS:The compound has melting point at 165-167°C. The result showed isolate was gartanin.CONCLUSION:The secondary metabolite found in pericarp Garcinia mangostana Linn. is gartanin.

Structural relationship among steroids from Sulawesi Tenggara’s sponge Clathria sp. and their radical scavenger activity

Four steroids were isolated and identified from the acetone extract of Clathria sp., namely clathruhoate or 3β-(butiryloxymethyl)-A-nor-5α-cholestane (1), 3β-(acetoxymethyl)-A-nor-5α-cholestane (2), 3β-(hydroxymethyl)-A-nor-5α-cholestane (3), and 3β-(hydroxymethyl)-A-nor-5α-cholest-15-ene (4). Isolation was carried out using chromatography techniques including Thin Layer Chromatography (TLC), vacuum liquid chromatography (VLC) and radial chromatography (RC) with silica gel as adsorbent and mixture of solvents as eluent. The Structure of isolated compounds was determined by spectroscopic methods i.e. FTIR, 1H and 13C NMR and also by comparing the spectroscopic data with similar data from references. The compounds isolated were evaluated for biological activity against a radical agent (DPPH radical). The relationship of the steroids showed that compound 3 is an oxidation product of compound 4. Clathruhoate (1) is formed by esterification of compound 3 with buthanoic acid, and compound 2 is produced by esterification of compound 3 with acetic acid. The radical scavenger potential of the methanol extract and steroids from Clathria sp. can be summarized as follows: methanol extracts > compound 4 > compound 3 > compound 2 > compound 1.

Opening of liver mitochondrial permeability transition pore in streptozotocin-induced diabetic rats and its inhibition by methanol fraction of Ficus mucoso (Welw) root bark

ObjectiveSeveral pathologies arise from the inappropriate opening of the mitochondrial permeability transition (mPT) pore. In this regard, inhibition of mPT pore represents a cytoprotective approach to preserve mitochondrial function for treatment of diseases characterized by excessive tissue wastage such as diabetes mellitus. The aim of this study, therefore, was to study the effects of fractions of Ficus mucoso, a medicinal plant used in the traditional treatment of diabetes, on mPT pore in normal and streptozotocin (STZ)-induced diabetic rat liver. MethodsDifferent solvent fractions of the crude methanol extract of F. mucoso were obtained by vacuum liquid chromatography and were tested on the mPT pore. Of all the fractions tested, methanol fraction of F. mucoso (MFFM) was the most potent and was used for in vivo studies. Diabetes mellitus was induced by a single intraperitoneal injection of 60 mg/kg STZ, while treatment lasted for 14 d. In vivo, 30 male Wistar rats were divided into five groups: A, normo-glycemic control (distilled water); B, STZ (65 mg/kg; diabetic control); C, STZ + MFFM (25 mg/kg); D, STZ + MFFM (50 mg/kg); E, STZ + glibenclamide (5 mg/kg). The mPT, mitochondrial ATPase activity, lipid peroxidation and cytochrome c release were assessed spectrophotometrically while blood glucose levels were monitored using glucometer. ResultsIn vitro, the solvent fractions of F. mucoso, at all concentrations tested, had no effect on the mPT pore, in the absence of calcium, with no significant release of cytochrome c. Interestingly, calcium-dependent pore opening was inhibited by all solvent fractions of F. mucoso, with the MFFM having the highest inhibitory effect of 83% at 3 mg/mL. Induction of opening of the mPT pore, significant (P < 0.001) enhancement of mitochondrial ATPase activity and elevated malondialdehyde (MDA) levels in STZ-induced diabetes were significantly (P < 0.001) reversed by MFFM and were comparable with the effects of glibenclamide, a standard antidiabetic drug. Also, treatment with MFFM at different doses significantly (P < 0.001) reduced high serum blood glucose compared to the diabetic control. ConclusionF. mucoso could be useful in therapeutic management of diabetes mellitus given its ability to prevent excessive tissue wastage via inhibition of pore opening, and reduction in levels of MDA and serum blood glucose.

Immunostimulant activity of steroid compound from the indonesian silver fern (P. calomelanos)

A secondary metabolite in steroid compound, namely β-sitosterol had been isolated from the n-hexane extract of the Indonesian silver fern. Extraction was conducted by maceration method, separation by chromatography techniques (vacuum liquid chromatography and flash chromatography), purification by recrystallization, and identification of its molecular structure by the spectroscopic methods and by comparison with reported literature data. The steroid isolate was yielded as colorless amorphous powder (melting point of 136– 137 °C). Based on the carbon clearance method, isolated steroid showed immunostimulant activity with phagocytosis index of 1.625.

CYTOTOXIC ACTIVITY OF ERIOCAULON CINEREUM R.BR TO MCF-7 AND VERO CELL LINE

Objective: The aim of the study was to determine the strength of the Eriocaulon cinereum R.Br plant against breast cancer cells (MCF7) and cytotoxicagainst Vero cells.Methods: E. cinereum R.Br was taken from the province of Bangka Belitung and then extracted in stages with n-hexane, ethyl acetate, and methanol.Then, the ethyl acetate extract was fractionated using the vacuum liquid chromatography method with dichloromethane and ethyl acetate solvents.Sample was tested for MCF-7 cells and Vero cells using the MTT method (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide). The dataobtained is analyzed by probit SPSS.Results: The results of this study showed that the best IC50 extract was ethyl acetate extract with an IC50 value of 450.31 μg/ml. Then, ethyl acetateextract produced dichloromethane fraction with IC50 value of 443.52 μg/ml and ethyl acetate with IC50 value of 214.75 μg/ml. Ethyl acetate fractionwas also tested against Vero cells to see cytotoxic against normal human cells with IC50 679.11 μg/ml