ANTIMICROBIAL CONSTITUENTS FROM INDIGENOUS PLANTS GROWING IN BANGLADESH

Abstract

This thesis report about the separation and structure explanation of secondary metabolite from the indigenous plant named Stereospermum suaveolens (Roxb.) DC(family-Bignoniaceae). A total eight (08) compounds were isolated of which one (SSC-003, Figure: 3.9) appears to be new. The compositions of the separated compounds were described mainly by NMR and mass spectroscopic techniques besides, Stereospermum suaveolens (Roxb.) DC and Diospyros malabarica (Desr.) Kostel (Family-Ebenaceae) buds was investigate for biological (antimicrobial, antioxidant, cytotoxic, thrombolytic, membrane stabilizing, analgesic, anti-diarrheal, CNS antidepressant and anti-diabetic) activity studies of the extractive from the plant. Repeated chromatographic isolation and purification of the n-hexane soluble part (HESF) of methanolic crude of the leaves and steam bark of S. suaveolens afforded two (02) compounds SSC-001 (Stigmasterol, Figure: 3.1) and SSC-002 (β-amyrin, Figure: 3.6). Between these two compounds SSC-002 (β-amyrin, Figure: 3.6) is separated for the first time from this plant. The carbon tetrachloride soluble part (CTSF) of leaves and steam bark of S. suaveolens yielded six (06) compounds designated as SSC-003 (19,23-dihydroxy-3-oxa-1-ursane, Figure: 3.9),SSC-004 (Uncaric acid, IUPAC name: 3β,6β,19α trihydroxyurs 12 ene 28 oic acid, Figure: 3.20), SSC-005 (Oleanolic acid Figure: 3.30), SSC-006 (p-Methoxy-trans-cinnamic acid, Figure: 3.35), SSC-007 (p-Methoxy-cis-cinnamic acid, Figure: 3.40) and SSC-008 (Ilexolic acid B, IUPAC name: 3β,19α dihydroxyolean 12 ene 23,28 dioic acid, Figure: 3.49). Compounds SSC-006 (p-Methoxy-trans-cinnamic acid, Figure: 3.35) and SSC-007 (p-Methoxy-cis-cinnamic acid, Figure: 3.40) have been also parted from chloroform soluble fraction (CLSF) of leaves and bark of S. suaveolens. Among these six compounds, SSC-003 (19,23-dihydroxy-3-oxa-1-ursane, Figure: 3.9) represents a 30-carbon containing triterpenoid which appears to be new. Rests of these five compounds are firstly isolated from this genus and species. The methanol extraction of S. suaveolens was researched for analgesic, antidiabetic, anti-diarrheal and anti-depressant activity on Swiss Albino mice. Antidiabetic activity was take measure by orally feed glucose tolerance test as the methanolic crude of S. suaveolens at 200- and 400- mg/kg b.w exhibited 37.58% and 56.10% decreased of blood glucose level as II along with 58.53% by standard glibenclamide (3 mg/kg b.w.) after 120 min induced the glucose. Castor oil-induced diarrhea in mice, the plant methanolic extract of 200- and 400- mg/kg b.w. demonstrated 23.08% and 46.15% while the standard loperamide (50 mg/kg b.w) exhibited 57.89% decrease of diarrheal feces. In tail flick method in mice for central analgesic activity, the methanolic extract dose of 200- and 400- mg/kg b.w. exhibited highest elongation (185.59% and 259.21% respectively) compared to 313.32% by morphine (4 mg/kg b.w) after 60 min. In peripheral analgesic activity assessment, the test samples, doses of 200- and 400- mg/kg b.w. displayed 31.82% and 48.48% reduction of writhing respectively as compared to 65.15% inhibition reduced by the standard diclofenac-Na (50 mg/kg b.w). In evaluation of anti-depressant activity, the crude of S. suaveolens (200- and 400-mg/kg b.w) didn’t show any remarkable CNS anti-depressant activity. The methanolic crude and its different Kupchan fragments of S. suaveolens were also based on screenings for total phenolic content, DPPH free radical scavenging assay, brine shrimp lethality bioassay (Cytotoxicity), membrane stabilizing, thrombolytic and antimicrobial activities. In antioxidant activity, aqueous soluble fraction of Kupchan partition showed highest IC50 value of 18.99μg/ml and highest phenolic content was found also (9.7 mg of GAE/gm of dried extract). Membrane stabilizing activity was assessed by hypotonic solution- and heat-induced methods and was compared to reference acetyl salicylic acid. In hypotonic solution-induced hemolysis, the n-hexane and the carbon tetrachloride soluble parts demonstrated 54.42% and 52.67% inhibition of hemolysis of RBCs. In heat-induced hemolysis, the chloroform soluble fragments revealed the hemolysis of RBC by 57.10% as compared to 72.09% produced by acetyl salicylic acid. In brine shrimp lethality bioassay, there was no remarkable lethality (IC50) observed. In the antimicrobial properties determination by disc diffusion method, only the n-hexane and carbon tetrachloride soluble parts revealed moderate activity (inhibition = 7.0-15.0 mm) against the test organisms. It is evaluated that the methanolic crude of S. suaveolens contains chemically unique and diversified chemical constituents. The plant extractives exhibited significant analgesic, anti-diarrheal and ant anti-hyperglycemic activities but moderate antioxidant, membrane stabilizing and thrombolytic properties. However, the test sample demonstrated weak antimicrobial activities. III The other part of present thesis was planned to assess the antioxidant, antimicrobial, antidiarrheal and analgesic activities of methanol crude and its different kapchan partitions of Diospyros malabarica (Desr.) Kostel buds growing in Bangladesh. In evaluation of antimicrobial assay, all the test samples showed prominent antimicrobial activity against the experimental organisms under in vitro conditions. Among these, the carbon tetrachloride soluble fragments exhibited the highest activity ranging 20-32mm in diameter. In antioxidant activity, aqueous soluble fraction of Kupchan partition showed highest IC50 value was 6.17 for DPPH assay test and also exhibited highest phenolic content was 12.87 mg of GAE/gm Gallic acid equivalence. In assessment of antidiarrheal activity, the D. malabarica extracts displayed significant anti-diarrheal potential in a dose dependent manner, the dose of 200- and 400- mg/kg b.w. displayed 26.92% and 50.00% while the standard loperamide (50 mg/kg b.w) revealed 53.85% reduction of diarrheal feces. During the assessment of analgesic activity by radiant heat tail-flick method, the plant sample at 400 mg/kg b.w. exhibited highest elongation (373.04%) as compared to morphine (435.88%) after 90 minutes of administration. In acetic acid-induced writhing test, the extract at 200- and 400-mg/kg b.w. showed 61.11 and 66.67% reduction of writhing in mice model, respectively as compared to 68.06% inhibited by the standard diclofenac-Na. The findings of this study justify some of the traditional uses of D. malabarica and reveal the bioactivity of the plants. Further studies are required to isolate and identify the bioactive compounds.