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Isolation, Structure Determination, and Antiaging Effects of 2,3-Pentanediol from Endophytic Fungus of Curcuma Amada and Docking Studies

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Isolation, Structure Determination, and Antiaging Effects of 2,3-Pentanediol from Endophytic Fungus of Curcuma Amada and Docking Studies Protoplasma (2014) 251:1089–1098 DOI 10.1007/s00709-014-0617-0 ORIGINAL ARTICLE Isolation, structure determination, and antiaging effects of 2,3-pentanediol from endophytic fungus of Curcuma amada and docking studies Sudeep Tiwari & Sailendra Singh & Pallavi Pandey & Shilpi K. Saikia & Arvind Singh Negi & Shailendra K. Gupta & Rakesh Pandey & Suchitra Banerjee Received: 22 November 2013 /Accepted: 16 January 2014 /Published online: 11 February 2014 # Springer-Verlag Wien 2014 Abstract An endophytic fungus was isolated from the rhi- (22.2 %) higher survival percentage was recorded in compar- zomes of Curcuma amada (Zingiberaceae), which was iden- ison to untreated control. The compound 1, however, lacked tified as Fusarium oxysporum on the basis of its morpholog- potential antimicrobial activity, indicating the plausible rami- ical and molecular characters. Chromatographic separation fication of the position of OH group in such bioactive mole- and spectroscopic analysis of the fungal metabolite (chloro- cules. In silico evaluation of these molecules against common form extract) led to the identification of one pure compound as well as unique targets corroborated better antiaging poten- having molecular formula C5H12O2, i.e., 2,3-pentanediol (1). tial of 1 in comparison to that of 2. The results for the first time Activity analysis of compound 1 demonstrated improved indicated that the utilization of the endophytic fungi of antiaging (antioxidant, thermotolerance) properties against C. amada could, thus, be a possible source for obtaining Caenorhabditis elegans, in comparison to a similar, commer- non-plant-based bioactive compounds having broader thera- cially available molecule i.e., 1,5-pentanediol (2). The effec- peutic applications pertaining to age-related progressions. tive (lower) concentration of 1 significantly showed (28.6 %) higher survival percentage of the worms under thermal stress Keywords Antiaging . Caenorhabditis elegans . Curcuma (37ºC) compared to its higher concentration (25.3 %), while amada . Endophytic fungus . Fusarium oxysporum . similar trends were followed in oxidative stress where 2,3-pentanediol . 1,5-pentanediol . Molecular docking . Oxidative stress Handling Editor: Jan Raoul De Mey Sudeep Tiwari and Sailendra Singh contributed equally to this work. Introduction S. Tiwari : S. K. Saikia : R. Pandey (*) Microbial Technology and Nematology Division, Central Institute of Endophytes are microorganisms that colonize intercellularly Medicinal and Aromatic Plants (CSIR-CIMAP), Lucknow, Uttar Pradesh 226015, India or intracellularly within healthy plant tissues and consequently e-mail: [email protected] represent a harmonious symbiotic relationship without caus- ing any apparent harm or disease symptoms to the host plants : : * S. Singh P. Pandey S. Banerjee ( ) (Chandra 2012). In recent years, fungal endophytes, especial- Plant Biotechnology Division, Central Institute of Medicinal and Aromatic Plants (CSIR-CIMAP), Lucknow, Uttar Pradesh 226015, ly those from medicinal plants, have drawn worldwide atten- India tion as they have not only mimicked the biosynthetic potential e-mail: [email protected] of their host plant but have also demonstrated potentials to synthesize unrelated bioactive molecules that find wide- A. S. Negi Medicinal Chemistry Division, Central Institute of Medicinal and ranging applications in medicine, agriculture, and industry Aromatic Plants (CSIR-CIMAP), Lucknow, Uttar Pradesh 226015, (Zhou et al. 2010;Linetal.2011; Bhagat et al. 2012; India Suryanarayanan et al. 2012). Advent of modern biotechno- logical tools has documented a far-reaching predictable prac- S. K. Gupta “ Department of Bioinformatics, Indian Institute of Toxicology and tical utility of economically important plant-inhabiting endo- Research (CSIR-IITR), Lucknow, Uttar Pradesh 226015, India phytic fungi” in industrial production of bioactive substances 1090 S. Tiwari et al. as part of their mutualistic relationships (Guo et al. 2008). in vivo bioactivity analysis of the fungal metabolite. Besides There is an increasing recognition of the vast repertoire of evaluating the antimicrobial property of the compound, anti- biomolecules produced by diverse group of fungal endo- oxidant and antiaging potentials of the same under thermal/ phytes, which can either be categorized as biologically active oxidative stress conditions had also been investigated and/or chemically novel molecules or can be broadly classi- employing a popular experimental model—Caenorhabditis fied as alkaloids, terpenoids, quinones, lignans, phenols, ste- elegans, which mimics the age-related physiological process- roids, isocoumarins, etc. (Lin et al. 2011; Bhagat et al. 2012). es of mammals, including humans (Guarente and Kenyon Numerous studies on isolation and categorization of endo- 2000; Wilson et al. 2006). phytic diversity from various medicinally important plant The present study reports Fusarium oxysporum as the species are progresssing, in order to unravel their relatively major endophytic fungi of C. amada through its isolation unstudied capability of synthesizing novel natural products for and molecular characterization, which yielded a distinctive therapeutic and industrial applications (Guo et al. 2008; fungal metabolite-2,3-pentanediol (1) that demonstrated both Bhagat et al. 2012;Chandra2012). It is however noteworthy in vivo and in silico antiaging potentials. that among the vast plant kingdom consisting of the nearly 300,000 plant species, only a handful of plant species has been studied with regard to their colonized endophytic populations, Materials and methods which leaves ample opportunity to find new and interesting endophytic microorganisms relating to the unstudied plant 1H, 13C and 2D nuclear magnetic resonance (NMR) spectra systems for isolating potential bioactive molecules similar to were recorded using Brukner Advance 300 DRX spectrometer that of the source plant and/or some “unnatural natural and the chemical shifts (δ) were expressed in ppm with products”or new metabolites having therapeutic/commercial reference to TMS as internal standard. ESI-MS data were potentials (Gunatilaka 2006; Aly et al. 2011; Bhagat et al. obtained in Shimadzu LC-MS system after dissolving com- 2012). pound in acetonitrile. Curcuma amada Roxb. (Family: Zingiberaceae), popularly known as mango ginger, is one such globally acclaimed Fungal isolation species of the genus Curcuma that stands next to turmeric in terms of its diverse healing properties owing to its potential to In vivo-grown C. amada rhizomes of single-plant sibling lines synthesize several major bioactive compounds, such as were collected from Chappra district of Bihar, India, and were curcumin, demethoxycurcumin, bisdemethoxycurcumin, di- planted at the research fields of CSIR-CIMAP (26°16′,N, terpene labda-8(17), and 12-diene-15,16-dial (Singh et al. 80°46′,E), Lucknow, India. After 2 months of acclimatization, 2010). Although several earlier studies have revealed percep- the fresh rhizome portions were aseptically collected. The tible bioactivities of the isolated endophytic assemblages of rhizomes were washed thoroughly under running tap water several other Curcuma species, namely Curcuma longa L. and subsequently surface sterilized with 70 % ethanol and (Maehara et al. 2011), Curcuma wenyujin (Wang et al. 0.1 % HgCl2 for 5 min, followed by 3–4 thorough washing 2012b), and Curcuma phaeocaulis (Bian et al. 2012), no with sterilized distilled water. The sterilized rhizomes were similar report is yet available with respect to the endophytes dried on sterile filter papers, dissected into small (3–5mm) of C. amada, which has been represented by a sole study pieces, and then placed on Potato Dextrose Agar (PDA) without any assigned bioactivity (Banu and Kumar 2009). medium supplemented with antibiotics (100 μg/ml streptomy- This information clearly indicates that functional identifica- cin) followed by incubation at 27±2 °C for 3–10 days. The tion of the most prevalent endophytic microorganisms of plates were methodically examined for fungal growth, and the C. amada coupled with activity-linked characterization of fungal hyphae spreading out from the explants were isolated the major chemical components of the same is the need of as single-fungal strain (coded as CIM-3) which was subse- the hour in view of their future applications. quently incubated on PDA for 7 days at 27±2 °C. Previous research indicated free-radical scavenging or an- tioxidant properties of C. amada metabolites (Policegoudra Fungal identification et al. 2007), but similar type of activity of the compound from the endophytic fungus of this plant is not yet explored. Genomic DNA of the exponentially growing culture of CIM-3 Moreover, many of the molecules of natural origin with anti- was extracted according to the manufacturer’s protocol using oxidant properties that scavenge ROS have attained a central the Zymo research (ZR) Fungal/Bacterial DNA MiniPrepTM stage in antiaging research (Huang et al. 2007; Shukla et al. Kit. Fungal 5.8s rDNA-ITS region was amplified from the 2012). In the background of this information, the present extracted genomic DNA by using the fungal domain specific study was designed to isolate, identify, and chemically char- primers of ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′)and acterize the major endophytic fungi of C. amada coupled with ITS2 (5′-GCTGCGTTCTTCATCGATGC-3′) (White et al. Microbe based antiaging potentials of 2,3-pentanediol 1091 1990). Each 50 μl of the PCR reaction
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