Journal of Pharmacy Research Vol.3.Issue 11.November 2010 Md. Abdul Muhit et al. / Journal of Pharmacy Research 2010, 3(11),2643-2646 2643-2646 Research Article ISSN: 0974-6943 Available online through www.jpronline.info *Corresponding author. Md. Abdul Muhit, Department of Pharmacy, University of Rajshahi, Rajshahi - 6205, Bangladesh, Tel: 0721-750041-54/4110, Fax: 0721-750064; E-mail: muhit_3533@yahoo.com INTRODUCTION Pterospermum acerifolium (Bengali name: Mushkand, Indian name: Kanak Champa, Muchukunda, family: Sterculiaceae) is extensively grown in North Canada, Bangladesh, Myanmar, several parts of India, sub-himalayan tracts etc. [1] In traditional system of medicines, The flower is sharply bitter, acrid; tonic, laxative, anthelmintic; removes “kapha”; inflammation, blood troubles, abdominal pain, ascites; cures ulcers, leprosy, urinary discharges, and tumors (Ayurveda). [2] The leaves are used as haemostatic and antimicrobial agent. [3,4] Previous investigations of bark of the plant have revealed the significant anti- ulcer [5] , anti-inflammatory, analgesic [6] , anti-oxidant activity [7] , and wound healing properties. [8] Previous Phytochemical studies showed the presence of acid polysaccharide [9] and flavonoids [10] in this species. Bark of the plant contains an acid polysaccharide composed of D-galacturonic acid, D-galac- tose, and L-rhamnose. Flowers are rich in carbohydrates with luteolin, kaempferol and quercetin. [1] Phytochemical and Biological Investigations of Pterospermum acerifolium Wild Bark. Md. Abdul Muhit *, Shamshad Shultana Khanam, Md. Saiful Islam, Mohammad S. Rahman, Bilkis Begum Department of Clinical Pharmacy and Pharmacology, Faculty of Pharmacy, University of Dhaka, Dhaka 1000, Bangladesh Received on: 15-06-2010; Revised on: 18-08-2010; Accepted on:13-09-2010 ABSTRACT Bioactivity guided phytochemical investigations of crude methanolic bark extract of Pterospermum acerifolium wild resulted in the isolation of three compounds for the first time such as (1) pentadec-11-enoic acid methyl ester, (2) oleanolic acid and (3) ß-sitosterol under silica gel CC and preparative TLC. The structures of the compound were elucidated by extensive spectroscopic studies ( 1 H NMR spectroscopy). The petroleum ether, carbon tetrachloride, dichloromethane and aqueous soluble partitionate of methanolic extract were subjected to antimicrobial, antioxidant and brine shrimp lethality bioassay. The average zones of inhibition produced by dichloromethane and carbon tetrachloride extract were found to be 13-16 mm and 12-13 mm respectively at a concentration of 400 μg/disc. In the brine shrimp lethality bioassay, LC 50 values obtained from the best fit line slope were 0.419, 1.362, 2.232, 1.867 and 2.874 μg/ml for Standard (Vincristine sulphate), petroleum ether, carbon tetrachloride, dichloromethane and aqueous soluble partitionate of methanolic extract respectively. All The four fractions showed significant antioxidant property using 1,1-diphenyl-2-pecrylhydrazyl (DPPH) scavenging assay along with total phenolic content, of which dichloromethane and aqueous soluble partitionate of methanolic extract demonstrated highest activity with IC 50 value of 26.5 and 39.0 μg/ml respectively. Key words: Pterospermum acerifolium , Kanak Champa, Antioxidant, Antimicrobial MATERIALS AND METHODS General experimental procedure Collection of plant materials P. acerifolium (Family: Sterculiaceae) wild bark was collected from Chittagong, Bangladesh in July 2007. The plant was identified and a voucher specimen (Accession number DACB 32446) representing this collection has been depos- ited in the Bangladesh National Herbarium, Dhaka, for further reference. Extraction and Isolation The stem bark of the plant was collected in fresh condition. The dried and coarse powder (700g) was extracted with methanol (2.5 liters) in an air tight, clean flat-bottomed container for 15 days at room temperature with occa- sional stirring. The extract was then filtered through a fresh cotton plug followed by a Whatman No. 1 filter paper. The filtrate was concentrated using a rotary evaporator at low temperature (40-45 o C) and pressure. The weight of the crude extract was 21.67 gm. Solvent–solvent partitioning was done using the protocol designed by Kupchan [17] and modified version of Wagenen et al. [18] The crude extract (5 gm) was dissolved in 10% aqueous methanol which was subsequently extracted first with petroleum ether (PE), then carbon tetrachlo- ride (CTC) and finally with dichloromethane (DCM). The aqueous methanolic extract was preserved as aqueous fraction (AQ). All the four fractions were evaporated to dryness by using rotary evaporator and kept in airtight contain- O O Compound 1(Pentadec-11-enoic acid methyl ester) 3 4 5 6 7 9 10 11 12 13 14 15 16 17 18 19 21 22 COOH H H HO 29 30 24 25 H 8 26 27 20 28 2 1 23 H 2 Compound 2 (Oleanolic Acid) The investigation was aimed to isolate the bioactive compounds and the scien- tific validation of the traditional uses of P. acerifolium bark against different Gram-positive, Gram-negative bacteria and fungi species using disc diffusion technique which probably most widely used of all other methods. [11] The cyto- toxic activity of the plant materials was performed by using brine shrimp lethality bioassay, which was proposed by Michael et al . [12] and modified by Solis et al . [13] is rapid, simple, inexpensive and requires small amount of test samples (2-10 mg or less). [14] The antioxidant property of the plants was evaluated using DPPH-free radical scavenging test, which was described previ- ously in several past decade. [15,16] The 1 H NMR spectra were recorded using an Ultra Shield Bruker DPX-400 (400 MHz) instrument. For NMR spectrum studies deuterated chloroform was used and the δ values for 1 H spectra were referred to the residual nondeuterated solvent signals. PTLC (20 x 20 cm) and TLC (20 x 5 cm) were carried out using Merck Si gel 60 PF 254 on glass plates at a thickness of 0.5 mm. Spots on TLC and PTLC plates were visualized by spraying the developed plates with vainillin-sulfuric acid followed by heating for 5 minutes at 110°c. All solvents used in this study were of reagent grade.