Document Type : Research Paper

Authors

1 M.Sc. Graduate of Medicinal and Aromatic Plants, Department of Agriculture, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, G. C., Tehran, Iran

2 Associate Professor, Department of Agriculture, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, G. C., Tehran, Iran

3 Assistant Professor, Department of Agriculture, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, G. C., Tehran, Iran

4 Assistant Professor, Department of Phytochemistry, Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, G. C., Tehran, Iran

Abstract

Abstract
Background and Objectives
Taxanes are a family of diterpene alkaloids which is currently known as the most important anticancer compounds to suppress and inhibit cell growth, differentiation and proliferation in indefinitely known cancer cell lines. The most well recognized member of the taxane family is taxol (paclitaxel), a minor cytotoxic component derived originally from yew bark (Taxus brevifolia). 10-Deacetylbaccatin III (10-DAB), which is a homolog of taxol, can be extracted at high yields from the needles of Taxus baccata and is currently considered the most appropriate precursor for semi-synthesizing taxol and its analogue, taxotere (docetaxel). The natural source of both compounds is the inner bark and needles of several Taxus species, especially T. baccata and T. brevifolia. Plant cell culture is an environmentally sustainable source of taxol and offers several advantages as it is not subjected to weather, season or contamination. The aim of the present study was to evaluate taxol and 10-DAB content of two Taxus species and their ability in production of both taxanes through cell culture.
Materials and Methods
The plant materials of T. baccata and T. berevifolia were collected from Shahid Beheshti University (35° 48' N, 51° 23' E at an altitude of 1785 m) and botanical garden of Faculty of Agriculture, University of Tehran, Karaj (35° 48' N, 50° 59' E at an altitude of 1238 m), respectively. Callus culture of both species was initiated from internodal segments on B5 medium supplemented with plant growth reaulators. Cell suspension culture was established on the medium supplemented with 2.0 mg L−1 naphtalene acetic acid (NAA) and 0.1 mg L−1 benzyl amino purine (BAP). Cell viability was determined by Evan’s blue staining test. After 21 days, the amount of taxol and 10-DAB was analysed by HPLC.
Results
The results showed that mother stocks of T. baccata and T. brevifolia contain 45 and 27 µg/g DW of taxol and 40 and 4 µg/g DW of 10-DAB, respectively. Cell growth was estimated by measuring fresh weight (FW) and dry weight (DW). Results showed that the maximum growth was reached after 21 days of cultivation. Cell cultures of T. baccata and T. brevifolia produced 540 and 20 µg/l taxol and 1 and 110 µg/l of 10-DAB, respectively. Taxol production in T. baccata cell culture was 26-fold higher than in T. berevifolia cell culture.
Discussion
As far as our literature survey could ascertain, growth and taxol production in T. baccata cell cultures have previously been reported. Although, T. baccata cells are more potent for the production of taxol, T. brevifolia cell culture can be considered for the production of 10-DAB, a homolog of taxol, as an appropriate precursor for semi-synthesizing taxol and its analogue, taxotere (docetaxel). However, it is essential to study and quantify the effect of selected key medium components on growth as well as product accumulation and strike a balance between the two to enhance the yield and productivity.

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References
Bedi, Y., Ogra, R., Koul, K., Kaul, B. and Kapil, R. (1996). Yew (Taxus spp.) A new look on utilization, cultivation and conservation. Supplement to cultivation and utilization of medicinal plants. Jammu-Tawi: Regional Research Laboratory.
Brunakova, K., Babincova, Z., Takac, M. and Cellarova, E. (2004). Selection of callus
cultures of Taxus baccata L. as a potential source of paclitaxel production. Engineering in Life Sciences. 4(5), 465-469.
Cusido, R.M., Palazon, J., Navia-Osorio, A., Mallol, A., Bonfill, M., Morales, C. and Pinol, M.T. (1999). Production of Taxol and baccatin III by a selected Taxus baccata callus line and its derived cell suspension culture. Plant Science, 146(2), 101-107.
Fornale, S., Degli Esposti, D., Navia-Osorio, A., Cusidò, R. M., Palazon, J., Pinol, M. T. and Bagni, N. (2002). Taxol transport in Taxus baccata cell suspension cultures. Plant Physiology and Biochemistry, 40(1), 81-88.
Gamborg, O. L., Miller, R. A. and Ojima, K. (1968). Nutrient requirements of suspension cultures of soybean root cells. Experimental Cell Research, 50(1), 151-158.
Ghassempour, A., Rezadoost, H., Ahmadi, M. and Aboul-Enein, H.Y. (2009). Seasons study of four important taxanes and purification of 10-deacetylbaccatin III from the needles of Taxus baccata L. by two-dimensional liquid chromatography. Journal of Liquid Chromatography & Related Technologies, 32(10), 1434-1447.
Green, F. J. (1990). The Sigma-Aldrich handbook of stains, dyes and indicators. . USA: Milwaukee, Aldrich Chemical Company Library.
Itokawa, H. and Lee, K. H. (2003) Taxus: The genus of Taxus. London: Taylor and Francis.
Kevin, W. and Rodney, C. (2001). Taxol biosynthetic genes. Phytochemistry, 58(1), 1-7.
Khosroushahi, A. Y., Valizadeh, M., Ghasempour, A., Khosrowshahli, M., Naghdibadi, H., Dadpour, M. R. and Omidi, Y. (2006). Improved Taxol production by combination of inducing factors in suspension cell culture of Taxus baccata. Cell Biology International, 30(3), 262-269.
Li, Y.C. and Tao, W. Y. (2009). Interactions of Taxol-producing endophytic fungus with its host (Taxus spp.) during Taxol accumulation.Cell Biology International,33(1), 106-112.
Nemeth-Kiss, V., Esther, F. and Tibor Cserh, G. (1995). Taxol content of various Taxus species in Hungary.Journal of Pharmaceutical and Biomedical Analysis, 14(8), 997-1001.
Nicolau, K. C., Yang, Z. and Liu, J. J. (1994). Total synthesis of taxol. Nature Biotechnology, 367(64), 630-634.
Onrubia, M., Moyano, E., Bonfill, M., Cusido, R. M., Goossens, A. and Palazon, J. (2013). Coronatine, a more powerful elicitor for inducing taxane biosynthesis in Taxus media cell cultures than methyl jasmonate. Plant physiology, 170(2), 211-219.
Palazon, J., Cusido, R.M., Bonfill, M., Morales, C. and Pinol, M. T. (2003). Inhibition of paclitaxel and baccatinIII accumulation by mevinolin and fosmidomycin in suspension cultures of Taxus baccata. Biotechnology, 101(2), 157-163.
Schippmann, U. (2001). Medicinal plants significant trade study. Germany: German Federal Agency for Nature Conservation Bonn.
Stierle, A., Strobel, G., Stierle, D., Grothaus, P. and Bignami, G. (1995). The search for a taxol-producing microorganism among the endophytic fungi of the pacific yew, Taxus brevifolia. Journal of Natural Products, 58(9), 1315-1324.
Tabata, H. (2004). Paclitaxel production by plant-cell-culture technology. Advances in Biochemical Engineering Biotechnology, 87, 1-23.
Vongpaseuth, K. and Roberts, S. C. (2007). Advancements in the understanding of Paclitaxel metabolism in tissue culture. Current Pharmaceutical Biotechnology, 8(4), 219-236.
Wani, M., Taylor, H., Wall, M., Coggon, P. and McPhail, A. (1971). The isolation and structure of taxol, a novel antileukemic and antitumor agentfrom Taxus brevifolia. American Chemical Society, 93(9), 23-25.
Zhong, J. J. (2002). Plant cell culture for production of paclitaxel and other taxanes. Journal of Bioscience and Bioengineering, 94(6), 591-599.