Crude extract of Trichoderma elicits agarwood substances in cell suspensionculture of the tropical tree, Aquilaria malaccensis Lam.

Agarwood is the precious fragrant wood produced by the tropical tree Aquilaria, often after elicitation by wounding or fungal attack. In this study we established a cell suspension culture of A. malaccensis from leaf-derived callus and induced agarwood production in the culture using fungal elicitors. Elicitors were made from crude mycelial extracts of two fungal species from the genera Trichoderma and Lasidiplodia. The elicitors were added to the cell suspension culture, initiated with 2 g of fresh calli, at concentrations ranging from 2 to 10 mg L-1. A light agarwood scent was detected from the suspension culture elicited with 8 mg L-1 Trichoderma extract. To increase scent intensity, cell suspension cultures were initiated from 2 to 8 g of calli and treated with 8 mg L-1 Trichoderma extract. The combination of 8 g of calli inoculum and 8 mg L-1 Trichoderma extract produced the most intense fragrance, one comparable to agarwood scent. The cell culture was harvested, extracted in methanol, and analyzed using GC-MS. Several important agarwood compounds were detected including 8-epi-.gama.-eudesmol, Ã¡-guaiene, and alloaromadendrene oxide-1. Trichoderma appeared to be a suitable inducer for agarwood production when used at an optimal concentration and in combination with a cell suspension culture of Aquilaria.

Anahtar Kelimeler:

Agarwood, cell culture, chemical constituents, endangered tree, GC-MS, Thymelaeaceae

Crude extract of Trichoderma elicits agarwood substances in cell suspensionculture of the tropical tree, Aquilaria malaccensis Lam.

Keywords:

Agarwood, cell culture, chemical constituents, endangered tree, GC-MS, Thymelaeaceae,

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Akalezi CO, Liu S, Li QS, Yu JT, Zhong JJ (1999). Combined effects of initial sucrose concentration and inoculum size on cell growth and ginseng saponin production by suspension cultures of Panax ginseng. Process Biochem 34: 639–642.
Barden A, Anak NA, Mulliken T, Song M (2000). Heart of the matter: agarwood use and trade and CITES implementation for Aquailaria malaccencis. Traffic Network Rep 46: 17–18.
Blackhall NW, Jotham JP, Azhakanandam K, Power JB, Lowe KC, Cocking EC, Davey MR (1999). Callus initiation, maintenance, and shoot induction in rice. In: Hall RD, editor. Plant Cell Culture Protocols. Vol. 111. Totowa, NJ, USA: Humana Press, Inc.
Bourgaud F, Gravot A, Milesi S, Gontier E (2001). Production of plant secondary metabolites: a historical perspective. Plant Sci 161: 839–851.
Chen H, Chen F (2000). Effects of yeast elicitor on the growth and secondary metabolism of a high-tanshinone-producing line of the Ti transformed Salvia miltiorrhiza cells in suspension culture. Process Biochem 35: 837–840.
Chen HQ, Wei JH, Yang JS, Zhang Z, Yang Y (2012). Chemical constituents of agarwood originating from the endemic genus Aquilaria plants. Chem Biodivers 9: 236–250.
Choi JW, Rim YK, Lee WH, Pedersen H, Chin CK (1999). Kinetic model of cell growth and secondary metabolite synthesis in plant cell culture of Thalictrum rugosum. Biotechnol Bioprocess Eng 4: 129–137.
CITES, Conventional on International Trade in Endangered Species of Wild Fauna and Flora (2012). Appendices I, II and III. Website: http://www.cites.org/eng/app/appendices.php [accessed 03 June 2013].
Guo S, Man S, Gao W, Liu H, Zhang L, Xiao P (2013). Production of ﬂavonoids and polysaccharide by adding elicitor in different cellular cultivation processes of Glycyrrhiza uralensis Fisch. Acta Physiol Plant 35: 679–686.
Jacinda TJ, Riaan M, Ian AD (2008). Characterization of two phenotypes of Centella asiatica in Southern Africa through the composition of four triterpenoids in callus, cell suspensions and leaves. Plant Cell, Tiss Org Cult 94: 91–99.
Jayaraman S, Daud NH, Halis R, Mohamed R (2014). Effects of plant growth regulators, carbon sources and pH values on callus induction in Aquilaria malaccensis leaf explants and characteristics of the resultant calli. J Forestry Res 25: 535–540.
Karam NS, Jawad FM, Arikat NA, Shibli RA (2003). Growth and rosmarinic acid accumulation in callus, cell suspension, and root cultures of wild Salvia fruticosa. Plant Cell, Tiss Org Cult 73: 117–121.
Kumeta Y, Ito M (2010). Characterization of delta-guaiene synthases from cultured cells of Aquilaria, responsible for the formation of the sesquiterpenes in agarwood. J Plant Physiol 154: 1998– 2007.
Lee CWT, Shuler ML (2000). The effect of inoculum density and conditioned medium on the production of ajmalicine and catharanthine from immobilized Catharanthus roseus cells. Biotechnol Bioeng 67: 61–71.
Leng TC, Ping NS, Lim BP, Keng CL (2011). Detection of bioactive compounds from Spilanthesacmella (L.) plants and its various in vitro culture products. J Med Plants Res 5: 371–378.
Mathur S, Shekhawat GS (2013). Establishment and characterization of Stevia rebaudiana (Bertoni) cell suspension culture: an in vitro approach for production of stevioside. Acta Physiol Plant 35: 931–939.
Mohamed R, Jong PL, Zali MS (2010). Fungal diversity in wounded stems of Aquilaria malaccensis. Fungal Divers 43: 67–74.
Mukherjee S, Ghosh B, Jha S (2000). Establishment of forskolin yielding transformed cell suspension cultures of Coleus forskohlii as controlled by different factors. J Biotechnol 76: 73–81.
Naef R (2011). The volatile and semi-volatile constituents of agarwood, the infected heartwood of Aquilaria species: a review. Flavour Frag J 26: 73–87.
Namdeo AG, Patil S, Fulzele DP (2002). Influence of fungal elicitors on production of ajmalicine by cell cultures of Catharanthus roseus. Biotechnol Prog 18: 159–162.
Okudera Y, Ito M (2009). Production of agarwood fragrant constituent in Aquilaria calli and cell suspension culture. Plant Biotechnol 26: 307–315.
Pornpunyapat J, Chetpattananondh P, Tongurai C (2010). Extraction kinetic model and chemical compounds of agarwood oil. In: Proceedings of the 2010 International Conference on Chemical Engineering and Applications (CCEA), pp. 321–326.
Saiful NT, Mashitah MY (2010). Chemical composition of volatile oils of Aquilaria malaccensis (Thymelaeaceae) from Malaysia. Nat Pro Comm 5: 1965–1968.
Savitha BC, Timmaraju R, Bhagyalaksami N, Ravishankar GA (2006). Different biotic and abiotic elicitors inﬂuence betalain production in hairy root cultures of Beta vulgaris in shake ﬂask and bioreactor. Process Biochem 41: 50–60.
Shrivastava N, Patel T, Srivastava A (2006). Biosynthesis potential of in vivo grown callus cells of Cassia senna I. var. senna. Curr Sci 90: 1472–1473.
Shu YQ, Meng LH, Li DL, Chuan HZ, Lan JH, Hui ZZ (2005). Production of 2-(2-phenylethyl)-chromones in cell suspension cultures of Aquilaria sinensis. Plant Cell, Tiss Org Cult 83: 217–221.
Simic SG, Tusevskli O, Antevskli S, Pancevska NA, Petreska J, Stefova M, Spasenoskli M (2012). Secondary metabolite production in Hypericum perforatum L. cell suspension culture upon elicitation with fungal mycelia from Aspergillus flavus. Arch Biol Sci Belgrade 64: 113–121.
Su WW, Lei F (1993). Rosmarinic acid production in perfused Anchusa officinalis culture: effect of inoculum size. Biotechnol Lett 15: 1035–1038.
Tabata Y, Widjaya E, Mulyaningsih T, Parman I, Wiriadinata H, Mandang YI, Itoh T (2003). Structural survey and artificial induction of aloeswood. Wood Res 90: 11–12.
Tamuli P, Boruah P, Nath SC, Leclercq P (2005). Essential oil of eaglewood tree: a product of pathogenesis. J Essent Oil Res 17: 601–604.
Thanh NT, Murthy HN, Yu KW, Jeong CS, Hahn EJ, Paek KY (2004). Effect of inoculum size on biomass accumulation and ginsenoide production by large scale cell suspension cultures of Panax ginseng. J Plant Biotechnol 6: 265–268.
Wang JW, Wu JY (2013). Effective elicitors and process strategies for enhancement of secondary metabolite production in hairy root cultures. Adv Biochem Engin/Biotechnol 134: 55–89.
Yang Y, He F, Yu L, Ji J, Wang Y (2009). Flavonoid accumulation in cell suspension cultures of Glycyrrhiza inflata Batal under optimizing conditions. Z Naturforsch 64: 68–72.
Yuan QS, Bi-Yao L, Liang-Feng Z (1992). Formation of oxo- agarospirol in Aquilaria sinensis. Plant Physiol Comm 28: 336–339.
Zhang CP, Li C, Yuan YJ, Sun AC, Hu CX (2001). Effects of fungal elicitor on cell status and taxol production in suspension cultures of Taxus chinensis var. mairei. Sheng Wu Gong Cheng Xue Bao 17: 436–440 (in Chinese).
Zhao J, Davis L, Verpoorte R (2005). Elicitor signal transduction leading to production of plant secondary metabolites. Biotechnol Adv 23: 283–333.
Zhong JJ, Yoshida T (1995). High-density cultivation of Perilla frutescens cell suspensions for anthocyanin production: effects of sucrose concentration and inoculum size. Enzym Microb Technol 17: 1073–1079.