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assess regulation of miRNA-mediated secondary metabolites biosynthesis in Swertia chir-

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metabolites in Swertia chirayita correlate with the expression proﬁles of multiple genes in

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plants of Swertia chirata Buch.-Ham. Ex Wall.: an endangered medicinal plant. Nat. Sci. 8

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servation implications. Plant Biosyst. 146 (2), 345e 351. Probatova, 2006. Chromosome numbers of plants of the primorsky territory, the Amur river

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chirayita. A Ph.D. thesis submitted to HNB Garhwal University Srinagar, Garhwal. Sharma, V., Barkha, K., Srivastava, N., Negi, Y., Dobriyal, A.K., Jadon, V.S., 2016. Assess-

ment of genetic ﬁdelity of in vitro raised plants in Swertia chirayita through ISSR,

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shoot multiplication and genetic validation in Swertia chirayita revealed through RAPD

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264690.

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Further reading

Davis, A.P., Govaerts, R., Bridson, D.M., Ruhsam, M., Moat, J., Brummitt, N.A., 2009.

A global assessment of distribution, diversity, endemism, and taxonomic effort in the

Rubiaceae. Ann. Mo. Bot. Gard. 96, 68e78.

242 CHAPTER 12 Swertia chirayita

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Jee, V., Dhar, U., Kachroo, P., 1983. Chromosome numbers of some alpine subalpine taxa of

Kashmir Himalaya. Herba Hung. 22, 23e31.

Mosaleeyanon, K., Zobayed, S., Afreen, F., Kozai, T., 2005. Relationships between net photo-

synthetic rate and secondary metabolite contents in St. John’s wort. Plant Sci. 169, 523e531.

Roy, S.C., Ghosh, S., Chatterjee, A., 1988. A cytological survey of eastern Himalayan plants.

II. Cell Chromosome Res. 11, 93e97.

Suryawanshi, S., Asthana, R.K., Gupta, R.C., 2009. Assessment of systemic interaction be-

tween Swertia chirata extract and its bioactive constituents in rabbits. Phytother Res. Int. J. Devot. Pharmacol. Toxicol. Eval. Nat. Prod. Deriv. 23 (7), 1036e1038.

Trillium govanianum

https://t.me/med1917

CHAPTER

Vishal Kumar1, Pradeep Singh2, Pramod Kumar Singh3, Mohammed Saba Rahim5,

Govt. Senior Secondary School, Bhadwar, Kangra, Himachal Pradesh, India;2Department of

Biotechnology, Guru Nanak Dev University, Amritsar, Punjab, India;

Christian Eminent College, Indore, Madhya Pradesh, India;

Bhag Singh University, Khiala, Jalandhar, Punjab, India;

Vikas Sharma

Agri-Food Biotechnology Institute, Mohali, Punjab, India

, Joy Roy5, Himanshu Sharma

Department of Biosciences,

Department of Botany, Sant Baba

Agri-Biotechnology Division, National

13.1 Introduction

Plants are major backbone to mankind in various ways. Since from early of civilization, people are dependent on natural resources and nature for various needs which we got from plant-based medicines/products as one of them. The Himalayan zone is harboring very rich diversity of ﬂora region. In India, the Himalayas are present in an area of w591,000 km E. Because of the Himalayas, India is considered in among 10 most comprehensively forested areas in the world and it is covered by 18% of India’s geographical area and subsequently forms more than 50% of the country’s forest cover and 40% of the species endemic to the Indian subcontinent (Saxena et al., 2002). Different climatic condi tions are mainly accountable for different types of diverse environments, which lead to higher biodiversity in the Himalayas. Rana and Samant

(2009) reported that there are more than 18,000 plants species that reside in the

Himalayas which include more than 1700 medicinally important plants (Samant

et al., 1998). Plants present in temperate and alpine climate are facing more stress

as compared to plants found in the subtropics, and due to this, they have unique medicinal and aromatic properties. Medicinal plants are mainly found in diverse habitats and habits, they may be annual, biennial, and others are perennial. Mankind has been using herbs and produce from different plants for healing ailments and improving well-being since antiquity.

Recent rises in cases of illness, side effects of allopathic medicines raises a concern on traditional medicines for various diseases and this enlightenment ultimately leads to the shift in the use of allopathic medicines to traditional medicines of plant origin. Several plants are used for the various medicinal purposes from immemorial times. Further plant-based medicines explained by various Unani, Ayurveda, and other medicine system impact a respectable position today, especially in the tribal regions of developing countries, where still health services are limited or

and lie between 27500and 37060N and 72300and 9725

Himalayan Medicinal Plants. https://doi.org/10.1016/B978-0-12-823151-7.00006-4

243

244 CHAPTER 13 Trillium govanianum

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not available for poor man. The popularity of traditional medicines which are more effective, safe, and inexpensive raised the concern among both developing and developed countries. Information of traditional medicine effectiveness has played a vital role in the discovery of novel products from plants as chemotherapeutic agents (Katewa et al., 2004). In addition to this, approximately 25% of pharmaceutical raw material comes directly from natural plant products (Schmidt et al., 2008). Continuous demands of traditional medicinal compound lead to the overexploitation of 1000 years conserved biodiversity. This leads to the creation of a situation where demand is huge and supply is lacking day by day. In most of the cases, demand is fulﬁlled by collecting material directly from the wild habitat which took very long time to revive and results into replenishment of that species. As already explained Himalayan region is one of the hotspots of biodiversity and also home for thousands of medicinal plant species. In the region of Himalayas, still 85% population relied on the traditional knowledge of medicinal plants (Farnsworth, 1988). Emphasis of the World Health Organization further puts pressure on this precious treasure of medicinal plants. Herbal industries are mainly dependent on its requirements on the medicinal plants of the Himalayan region (Dhar et al., 2000). Continuous widening of gaps between demand and supply leads to illegal harvesting which in result puts threat to many species at the brink of extinction (Vidyarthi et al., 2013). Among the medicinal plants present in Himalayan region, Trillium govanianum is one of the fast-emerging medicinal herbs, with many pharmacological activities.

13.1.1 Trillium govanianum

T. govanianum Wall ex D. Don, commonly known as Himalayan Trillium, Nag chhatri, or Teen patra, is a perennial herb endemic to the Himalayas (Samant et al., 1998;

Kubota et al., 2006). It belongs to the plant family Melanthiaceae, having 181

species related to 17 genera of perennial herbs. Mostly distributed in the temperate region of Northern Hemisphere and considered for its traditional and modern medicinal properties (Zomlefer et al., 2001; Christe nhusz and Byng, 2016), T. govanianum belongs to genus Trillium, one of the largest genus of Melanthiaceae, comprising of 50 species, 39 American species of Arcto-Tertiary origin, and 11 Asian species (http://www.theplantlist.org/1.1/browse/A/Melanthiaceae/). This genus is further divided into two subgenera, Phyllantherum Raf. comprising of sensile-ﬂowered species (26 American species) and Trillium with pedicellate-ﬂowered species (all Asian and remaining American species) (Ohara and Kawano, 2006; Schilling

et al., 2019 ). Only two species, namely, i.e., T. govanianum and Trillium tschonoskii

of genus Trillium are found in the Himalayan region.

T. govanianum has an important place in the traditional medicine system due to the presence of various therapeutically important compounds in its rhizomes (Zhan, 1994; Shah, 2006; Khan et al., 2016). Recently, a trend rises in the collection of T. govanianum from the forests attracts the attention of biodiversity conservators and researchers. The important part for which it is exploited is undergrou nd

13.2 Classiﬁcation, origin, distribution, and cytotaxonomy 245

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rhizome containing trillarin, a r aw material for medicines mainly sex stimulants. In traditional medicines system, rhizom e of T. govanianum is used for the cure of dysentery, boils, menstrual and sexual disorders, etc. The recent studies led to the identiﬁcation of several p h a r m a co logical activities possessed by T. govanianum such as antiseptic, analgesic, antiinﬂammatory, antifungal, free radical scavenging, as well as cytotoxicity against prostate and cervical carcinoma cells (Ur Rahman

et al., 2016).

Recently, important active components are explored and their corresponding pharmacological activities from Trillium are of utmost importance. It is speciﬁcally distributed in the Indian Himalayas, and collecting its rhizome for commercial activities has become common in the Indian Himalayas. However, in reality, the collection of species from natural habitats is unjustiﬁable. Understanding the phenomenon like socioecological dynamics of a species and making a strategy for its sustainable use is a challenging mission in the Himalayas. There should be potential and effective strategies for conservation of the species while retaining community incomes. This chapter provides a focus on overview of the biology, uses, and conservation approaches that can be followed for the sustainable utilization of T. govanianum in the Indian Himalayas.

13.2 Classiﬁcation, origin, distribution, and cytotaxonomy

13.2.1 Classiﬁcation and morphology

Kingdom: Plantae Subkingdom: Tracheobionta Superdivision: Spermatophyta Division: Magnoliophyta Class: Liliopsida Subclass: Liliidae Order: Liliales Family: Melanthiaceae Genus: Trillium L. Species: govanianum

T. govanianum is a small 100e200 mm creeping stem rising from a short, tuber- ous rhizome and having one or three ovate, acute, stalked leaves. At the time of reproduction, a ﬂower emerges at the shoot apex, surrounded by three leaves. Flowers are 2e3 cm long having six distinctly yellow stamens ﬁlament attached to base, a whorl each of petals and sepals, and a three-celled purplish brown ovary that produces multiple seeds. The fruit is a globular red berry and seeds are ovoid with pulpy lateral appendages (Fig. 13.1). Seeds of Trillium spp. are numerous and their dispersal is low, such that seeds typically remain close to the parent plant (Ohara and Kawano, 2005).

246 CHAPTER 13 Trillium govanianum

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FIGURE 13.1

Trillium govanianum plant growing in wild habitat.

13.2.2 Distribution

This plant is mainly distributed in the range of 2500e4000 mts across the Himalayas (Vidyarthi et al., 2013). The plant is sciophyte having three leaves on purple red stem bearing trimerous ﬂowers. T. govanianum is most commonly found under the canopies of mix temperate (Abies pindrow, Betula utilis, Cedrus deodara, Juglans regia, Juniperus indica, Picea smithiana, Quercus spp., Rhododendron spp., and Salix spp.) and subalpine forests (Rhododendron spp.) with habitat of thick humus decomposing litter. Due to these speciﬁc habitat conditions, the species has patchy and limited distribution to speciﬁc pockets in the Himalayas (Chauhan et al., 2020). The species is more common in western regions of the Himalayas as compared to eastern region (Chauhan et al., 2018). The main areas of its distribution are as follows: in Jammu and Kashmir, the species grows in Fatehpur, Gulmarg, Gurez, Kanzalwan, Pahalgam, Poonch, Sonamarg in Uttarakhand, Gangotri, Govind Pashu Vihar, Harshil, Kedarnath, Munsiyari, Panchachuli, Pindari, Sunderdhunga, and Tungnath, while in areas of Himachal Pradesh, the species is commonly reported from Kullu, Kinnaur, Lahaul-Spiti, and Shimla.

13.2.3 Origin and cytotaxonomy

With a base number (x ¼ 5), genus Trillium possesses different ploidy levels. Among the Asian species, except Trillium camschatcense (diploid, 2n ¼ 10) all the species are allopolyploids involving the hybridization of different genomes, whereas American species are diploid 2n ¼ 10, except for few reports of autotriploids (Darlington and Wylie, 1956; Darlington and Shaw, 1959). The karyological analysis of T. govanianum has revealed that it is a tetraploid, 2n ¼ 20, having 10 pairs (AeJ) of unusual long chromosomes. Based on length and position of centromere, chromosome pairs are categorized as (i) metacentricesubmetacentric: ﬁrst two large pairs (A and B), (ii) submetacentric: two next large (C and D) and two small (I and J), and (iii) acrocentric: four moderate sized (E, F, G, and H) (Mehra

and Sachdeva, 1975).

13.3 Biochemical analysis 247

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Based on cytological and morphological characteristics, the origin of T. govanianum is considered as a historic evolutionary event for survival and perpetuation against several cold and dry seasons during Pleistocene age in Asia.

T. govanianum originates from the rare natural intergeneric hybridization of genus Trillium and Daiswa (Fukuda, 2001).

13.3 Biochemical analysis

The genus Trillium having around 31 species and few of them consists of rich sources of bioactive compounds which possess medicinal value with wide application in pharmaceuticals application (Gracie and Lamont, 2012). The major classes of phytochemicals include mainly steroids, glycosides, terpenoids, sterol, saponins, and ﬂavonoids (Ismail et al., 2015). Various protocols have been reported for efﬁcient extraction of metabolite from dried rhizomes of T. govanianum and characterized using various chromatographic techniques such as NOESY, HSQC, HMBC, FAB, HR-FAB, IR, and UV. Previously only four com pounds, namely Govanoside A, Pennogenin, Borossoside E and Diosgenin were detected. Recently, Singh et al. (2020) also identiﬁed 24 steroidal saponins in T. govanianum by using UHPLC-QTOF-MS/MS (Fig. 13.2).

H-NMR,13C-NMR, COSY,

FIGURE 13.2

Biochemical structure of phytochemicals present in Trillium govanianum.

Based on Ur Rahman, S., Adhikari, A., Ismail, M., Shah, M.R., Khurram, M., Anis, I., Ali, F., 2017a. A new

trihydroxylated fatty acid and phytoecdysteroids from rhizomes of Trillium govanianum. Record Nat Prod 11,

323e327; Ur Rahman, S., Ismail, M., Khurram, M., Ullah, I., Rabbi, F., Iriti, M., 2017b. Bioactive steroids and

saponins of the genus Trillium. Molecules 22(12), 2156.

248 CHAPTER 13 Trillium govanianum

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Table 13.1 List of bioactive compounds in the Trillium species.

S. no. Source Chemical compound Class References

1 Trillium

govanianum

2 T. govanianum Borossoside E Sterol saponin Ur Rahman

3 T. govanianum Pennogenin Sterol saponin Ur Rahman

4 T. govanianum Diosgenin Sterol saponin Ur Rahman

5 T. govanianum Govanoside A,

Govanoside A Sterol saponin Ur Rahman

Govanoside B, Protodioscin, Pregnachacotrioside, Pennogenin tetraglycosides, Pennogenin diglycosides, Borassoside E, Borassoside D, Diosgenin, Pennogenin-

D-glucopyranosyl-

[O-bS1 or its isomer, (1b, 23S,24S)-1-[O-bglucopyranosyl (1 / 3)-

D-xylopyranosyl-(1

O-(b/ 2)-O-a-

L-

rhamnopyranosyl]-23 hydroxyspirosta5,25-dienyl-24-[O-b-D6-deoxygulopyranoside] or its isomer, pirosta5,25-dienyl-[O-bglucopyranosyl-S3 or its isomer, Protodioscin, Pennogenin-[O-bglucopyranosyl-S4 or its isomer, Diosgenin-[O-b-

D-glucopyranosyl-S5 or

its isomer, Pennogenin-

D-glucopyranosyl-

[O-bS4 or its isomer, Pennogenin-[O-bglucopyranosyl-S6, Gentrogenin 3-O-bchacotrioside or its isomer, Pennogenin-

D-glucopyranosyl-

[O-b-

et al. (2015a,b)

Sterol saponin Singh et al.

(2020)

D-

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