RESEARCH PAPER
Remedial effect of ascorbic acid and citric acid on oxidative browning of Glycyrrhiza glabra callus cultures
More details
Hide details
1
Department of Biotechnology, Faculty of Engineering and Technology,
Manav Rachna International University, Faridabad, India
Submission date: 2016-06-09
Final revision date: 2016-07-18
Acceptance date: 2016-07-26
Publication date: 2016-11-02
BioTechnologia 2016;97(3):179-186
KEYWORDS
TOPICS
ABSTRACT
Oxidative browning is a common and severe problem in plant tissue cultures including callus cultures of Glycyrrhiza glabra , where browning is caused by accumulation and oxidation of phenolic compounds. Brown callus results in decreased biosynthetic potential and poor growth, eventually leading to cell death. In this study, the effect of different concentrations and combinations of antioxidants – ascorbic acid, and citric acid on browning of G.glabra callus was studied. Morphological and ultra-structural changes related to browning and antioxidant treatment were also studied using scanning electron microscopy (SEM) and transmission electron microscopy (TEM) to evaluate the cellular characteristics and metabolic activity of control and treated callus. Intensity of browning in callus remarkably decreased with increasing concentrations of ascorbic acid; the browning was completely overcome at a concentration of 20 mg/l ascorbic acid, in combination with 10 mg/l citric acid. An electron microscopy analysis showed that brown callus was found to have ruptured cell surface and a disorganized cell structure with the absence of cell organelles that affected callus development and lead to cell death. Non-brown callus had all the characteristics typical for a non-embryogenic callus cells with high-level activity showing all vital cell organelles including abundant and elongated mitochondria. Therefore, we conclude that ascorbic acid and citric acid have wide applications in tissue cultures of Glycyrrhiza glabra where browning restricts the callus development, and lead to the establishment of non-embryogenic callus with high metabolic activity for further use in secondary metabolite production.
REFERENCES (32)
1.
Aslam J., Khan S. A., Cheruth A. J., Mujib A., Sharma M. P., Srivastava P.S. (2011) Somatic embryogenesis, scanning electron microscopy, histology and biochemical analysis at different developing stages of embryogenesis in six date palm (Phoenix dactylifera L.) cultivars. Saudi J. Biol. Sci. 18: 369-380.
2.
Banerjee S., Upadhyay N., Kukreja A.K., Ahuja P.S., Kumar S., Saha G.C. (1996) Taxanes from in vitro cultures of the Himalayan yew Taxuswallichiana. Planta Medica 62: 329-331.
3.
Bradford M.M. (1976) A rapid and sensitive method for quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 72: 248-254.
4.
Canhoto J.M., Mesquita J.F., Cruz G.S. (1996) Ultrastructural changes in cotyledons of pineapple guava (Myrtaceae) during somatic embryogenesis. Ann. Bot. 78: 513-521.
5.
Chaudhury A., Qu R. (2000) Somatic embryogenesis and plant regeneration of turf-type bermudagrass: effect of 6- benzyladenine in callus induction medium. Plant Cell Tiss Org. 60: 113-120.
6.
Dehon L., Macheix J.J., Durand M. (2002) Involvement of peroxidases in the formation of the brown coloration of heartwood in Juglans nigra. J. Exp. Bot. 53: 303-311.
7.
Figueiredo M.A., Paiva R., Souza A.C., Porto J.M.P., Nogueira G.F., Soares F.P. (2007) Indução in vitro de calosemduase spécies de maracujazeironativo. Rev. Brasil. Biociên. 5: 288-290.
8.
Flora S.J.S. (2009) Structural, chemical and biological aspects of antioxidants for strategies against metal and metalloid exposure. Oxid. Med. Cell Longev. 2: 191-206.
9.
Fu C., Lei C., Gan L., Li M., Yang Y., Yu L. (2010) Optimization of embryogenic-callus induction and embryogenesis of Glycyrrhiza glabra. Afr. J. Biotechnol. 9: 5823-5829.
10.
Habibi N., Suthar R.K., Purohit S.D. (2009) Role of PGrs and inhibitors in induction and control of somatic embryogenesis in Themedaquadrivalvis. Indian J. Exp. Biol. 47: 198-203.
11.
Havsteen B.H. (2002) The biochemistry and medical significance of the flavonoids. Pharmacol. Ther. 96: 67-202.
12.
He Y., Guo X., Lu R., Niu B., Pasapula V., Hou P., Cai F., Xu Y., Chen F. (2009) Changes in morphology and biochemical indices in browning callus derived from Jatropha curcas hypocotyls. Plant Cell Tiss. Organ. Cult. 9: 11-17.
13.
Hossain M.J., Bari M.A., Ara N.A., Shahinulislam S.M. (2009) Effect of carbon sources on cell growth and regeneration ability in three cultivars of banana. J. Biosci. 17: 83-88.
14.
Huo Y.Q., Ge S.J., Men Y.J., Liu G.R. (2005) The progress of tissue culture of the Chinese liquorice (Glycyrrhiza uralensis Fisch.). Chin. Agric. Sci. Bull. 21: 64-66.
15.
Jedinak A., Farao J., Psenakova I., Maliar T. (2004) Approaches to flavonoid production in plant tissue cultures. Biologia 59: 697-710.
16.
Khosroushahi A.Y., Manesh H.N., Simonsen H.T. (2011) Effect of antioxidants and carbohydrates in callus cultures of Taxusbrevifolia: evaluation of browning, callus growth, total phenolics and paclitaxel production. Bioimpacts 1: 37-45.
17.
Laukkanen H., Häggman H., Kontunen-Soppela S., Hohtola A. (1999) Tissue browning of in vitro cultures of Scots pine: role of peroxidase and polyphenol oxidase. Physiol. Plant. 106: 337-343.
18.
Mayer A., Harel E., Shaul R. (1965) Assay of catechol oxidase a critical comparison of methods. Physicochemistry 5: 783-789.
19.
Mousa N., Siaguru P., Wiryowidagdo S., Wagih M.E (2006). Rapid clonal propagation of Licorice (Glycyrrhiza glabra) by in vitro. Sugar Technol. 8: 292-298.
20.
Murashige T., Skoog F. (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15: 473-497.
21.
Nakamura T., Taniguchi T., Maeda E. (1994) Cyto-histological studies on somatic embryos of Coffee: ultrastructural aspects. Japan. J. Crop Sci. 63: 144-157.
22.
Ribeiro O., Paiva L.V., Pádua M.S., Santos B.R., Alves E., Stein V.C. (2012) Morphological and ultrastructural analysis of various types of banana callus, cv. Prata ana. Acta Sci. 34: 423-429.
23.
Rocha D.I., Vieira L.M., Otoni W.C., Tanaka F.A.O., Da Silva L.C.(2012) Somatic embryogenesis of a wild passion fruit species Passiflora cincinnata Masters: Histological and histochemical evidences. Protoplasma 249: 747-758.
24.
Sawaengsak W., Saisavoey T., Chuntaratin P., Karnchanatat A. (2011) Micropropagation of the medicinal herb Glycyrrhiza glabra L. through shoot tip explant culture and glycyrrhizin detection. Int. Res. J. Plant Sci. 2: 129-136.
25.
Sharma P., Uppal S., Sehrawat A.R. (2005) Evaluation of callus cultures of Glycyrrhiza glabra L. var. HM 1 for glycyrrhizin. Nat. J. Plant Improv. 7: 62-64.
26.
Steinmacher D.A., Guerra M.P., Saare-Surminski K., Leiberei R. (2011) A temporary immersion system improves in vitro regeneration of peach palm through secondary somatic embryogenesis. Ann. Bot. 108: 1-13.
27.
Uddain J., Gnasekaran P., Zakaria L, Lynn C.B., Subramaniam S. (2015) The Effect of different growth media, Carbon source and PGRs on Dendrobium broga giant orchid’s protocorm-like bodies (plbs) proliferation supported with SEM and TEM analysis. Pak. J. Bot. 47: 587-593.
28.
Vijayalakshmi U., Shourie A. (2013) Gas Chromatography- Mass spectrometric analysis of Glycyrrhiza glabra Linn. Roots. Int. J. Pharm. BioSci. 4: 741-755.
29.
Vijayalakshmi U., Shourie A. (2015) Elicitor induced flavonoid production in callus cultures of Glycyrrhiza glabra and regulation of genes encoding enzymes of the phenyl propanoid pathway. Der Pharmacia Lett. 7: 156-166.
30.
Wang M., Liao F., Yang L, Huang D.L., Yang L.T., Li Y.R. (2016) Influence factors and cell structure changes related to sugarcane stem tip browning in vitro culture. IJAIR 4: 768-772.
31.
Wongwichaa W., Tanakab H., Shoyamac Y., Tuvshintogtokhd I., Putaluna W. (2008) Production of Glycyrrhizin in callus cultures of licorice journal of biosciences. Z. Naturforsch. C 63: 413-417.
32.
Yu L.Q., He M.T., Wang Z.L., Xu Z.Y (1999) Study on techniques for rapid propagation of licorice (Glycyrrhiza uralensis Fiseh) by tissue culture. Grassland China 1: 12-14.