TLC and GC-MS analysis of fermented wood "Nikhra

10.31195/ejejfs.417112 Eurasian Journal of Forest Science 2018 6(3): 1-7 http://dergipark.gov.tr/ejejfs TLC and GC-MS analysis of fermented wood "Nikhra" petroleum ether fraction of Combretaceae spp. Combretum hartmannianum and Terminalia laxiflora Noha Fadle1, Abdalbasit Adam Mariod2*, Hiba Abdel Rahman Ali3 , Alfatih Ahmed Hasan4 1 The National Centre for Research, Environment and Natural Resources Research Institute, Khartoum, Sudan 2* Indigenous Knowledge Center, Ghibaish College of Science and Technology, Ghibaish, Sudan. 3 Commission for Biotechnology and Genetic Engineering (CGEB), National Centre for Research 4 Department of Organic Chemistry, College of Science, Sudan University of Science and Technology, P.O. Box , Khartoum, Sudan. Corresponding author: , Abstract This study aims to analysis fractions (petroleum ether, chloroform, methanol and aqueous) fermented wood Nikhra of Combretaceae spp (Combretum hartmannianum and Terminalia laxiflora) analysis it by using chromatographic and spectroscopic analysis. Petroleum ether Nikhra fraction analysis with TLC and spray TLC with vanillin H2SO4 (pink) (B1, B2), Rf values (0.84, 0.81), were expected to be phenolic, with vanillin HCL (red) compounds spots (B2) with Rf values (0.81) was expected to be catechin and with vanillin H3PO4, blue-violet zones compounds spots (A1, A2, A3), (B1, B2, B3, B4), with Rf values (0.88, 0.78, 0.67), (0.84, 0.81, 0.67, 0.59), respectively were expected to be lignans. Petroleum ether fermented wood"Nikhra" fraction was divided into two types of compounds classes aromatic and non-aromatic and hence compounds were classified to phenolics and terpenoids compounds by GC/MS. Fragrant aromatics or terpenoids were targeted in this part of study. GC-MS analysis gave a spectrum of fragrance aromatic compounds (phenolics) in the petroleum ether Nikhrafractions of T. laxiflora, was Lup-20(29)-en-3-ol, acetate, (3β) and Tetracosamethylcyclododecasiloxane, main terponoids compounds were eicosamethylcyclodecasiloxane. Main fragrance aromatics compound in the petroleum ether Nikhra fraction of C. hartmannianum was 2-tert-Butyl-5(hydroxtmethyl)-4-formylfuran, and main terponoids compound was Tetracosamethylcyclododecasiloxane. Key words: Combretaceae, Combretum hartmannianum, Terminalia laxiflora, Gas Chromatography, Thinlayer chromatography (TLC) Introduction The resinous heartwood of C. hartmannianum and T. laxiflora, trees are usually used in Sudanese fragrances, the scented heart wood is used as perfume and the root bark is used to treat wound and strains, the macerated stem bark of T. laxiflora serves as antiseptic to wash mouth in order to resist gingivitis and thrush serves as wound dressing, diuretic management, pile and yaws treatment(Ivory coast), anti-skin inflammation, sores and ulcers treatment (Sierra Leone), eye lotion(Gambia), hair perfume, severe jaundice and chewing stick (Cameroon) across other African 1 Eurasian Journal of Forest Science - TLC and GC-MS analysis of fermented wood “Nikhra” by Fadle et al.2018 countries (Abbiw, 1990; Daniel, 1990; Batawila et al., 2005). The barks decoction of T. laxiflora is used for malaria (Doka and Yagi, 2009). Dokhan and Bakhour are a traditional processes used by Sudanese married women to make their own perfume, rarely single female use them for medical purpose (Mariod et al., 2014). The present study, is designed to analyze fractions (petroleum ether, chloroform, methanol and aqueous) of fermented wood Nikhra-Combretaceae spp (Combretum hartmannianum and Terminalia laxiflora) by using chromatographic and spectroscopic analysis. Materials and Methods Collection of plant materials Natural fermented hardwood "Nikhra" of Combretum hartmannianum and Terminalia laxiflora were collected from Kordofan state, Sudan. They were, carefully, washed, oven-dried for 1 h at 50°C and ground into a fine powder. Plant materials preparation and extraction 100 g of ground powder of each plant Nikhra were extracted by Soxhlet apparatus using methanol. The methanolic extract was fractionated, sequentially, using solvents of increasing polarity namely petroleum ether, chloroform, and aqueous. Fractions were dried using a rotary evaporator and stored at 4°C for further analysis (Fyhrquist et al., 2002). Thin Layer Chromatographic (TLC) analysis TLC was performed on a pre-coated silica gel TLC plates grade F254 (E-Merck, Darmstadt, Germany) to determine the number of compounds present in petroleum ether Nikhra fraction. Sample was spotted at 1.0 cm from the bottom of silica gel plates using capillary tubes. Development of the chromatogram was done in closed tanks, in which the atmosphere has been saturated with eluent vapor by wetting a filter paper lining. The chromatogram was visualized under UV light (366 nm and 254 nm), Natural Product Reagent (NPR), H2SO4, HCL and H3PO4 acid reagent spray. The Rf values of the compounds were calculated using the following formula. Rf = distance travelled by the compound/Distance travelled by the solvent front GC-MS Analysis Petroleum ether Nikhra fraction was subjected to GC MS analysis to identify the various bioactive compounds present. The sample was analyzed in GCMS- QP2010 Plus from Delhi University (DU) India. ACQ Mode Scan: 40m/z to 600m/z, Column flow is 1.21mL/min and total flow is 16.3ml/min. Flow control with linear velocity is 40.9cm/sec. The identification of compounds was done using computer matching of mass spectra with those of standards (WILEY8. LIB. and NIST11.library). Results and Discussion Thin layer chromatographic analysis The presence of flavonoids was confirmed by their color change from quenching fluorescence (254nm) to yellow or orange color for flavonoid and prominent blue color in case of flavonoidal acids or other phenolic acids (366 nm) after spraying with Natural Product Reagent (NPR) (Table 1). Polyphenols (phenolic acids, flavanoids) has been detected using NPR (366 nm). Fluorescence 2 Eurasian Journal of Forest Science - TLC and GC-MS analysis of fermented wood “Nikhra” by Fadle et al.2018 behavior of flavonoids in response to NPR is structure dependent. Flavonoids e.g. quercetin and myrecitin develops orange color and those of kaempferol and isorhamntin yellow to green colors. Flavones glycosides of luteolin develops orange colors and those apigenin yellow to green (Wagner and Bladt, 1996). Vanillin H2SO4 is a universal reagent that detects components of the petroleum ether fraction, terpenoids, phenols etc., typical pink to purple colors were developed upon spraying with vanillin H2SO4 (heat 110ᵒ C). All phenolic at UV 254 nm show prominent quenching, and they give blue fluorescence at UV 366 nm (Wagner and Bladt, 1996). After spraying fraction of the petroleum ether of fermented wood of T. laxiflora and C. hartmannianum by vanillin H2SO4, they showed typical pink and purple zones of phenolic. Accordingly compounds spots (B1, B2), Rf values (0.84, 0.81), were expected to be pheno (...truncated)