Pharmacognostical studies of Cleome viscosa Linn.

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1 Indian Journal of Natural Products and Resources Vol. 3(4), December 2012, pp Pharmacognostical studies of Cleome viscosa Linn. Prakash Chandra Gupta 1, 2 * and Ch V Rao 3 1 University Institute of Pharmacy, Chhatrapati Shahu Ji Maharaj University, Kanpur , Uttar Pradesh, India 2 Institute of Pharmaceutical Sciences and Research Centre, Bhagwant University, Ajmer , Rajasthan 3 Pharmacognosy and Ethnopharmacology Division, National Botanical Research Institute, Lucknow , Uttar Pradesh Received 4 October 2011; Accepted 14 May 2012 Cleome viscosa Linn. (Family: Capparaceae) is an annual, sticky herb and commonly known as wild or dog mustard. It has various ethnomedicinal values as various traditional communities find diverse medicinal properties. This work aimed to study the macro- and microscopical, physicochemical, phytochemical and fluorescence analysis of different parts of the C. viscosa. Macro- and microscopical studies showed the presence of palmately compound leaf with 3-5 leaflets, obovate to lanceolate, glandular and simple covering trichomes and anomocytic arrangement of stomata. Sclerenchmatous caped vascular bundle in petiole, arc shaped pericyclic fibres in stem and distinct uni to biseriate medullary rays of root are some of the diagnostic features noted from anatomical study of the plant. Powder microscopy revealed the presence of fibres, spiral and pitted vessels, trichomes and calcium oxalate crystal. Total ash of leaf was about two times higher than root and about four times higher than seed. The fluorescent analysis under day light and UV light by treatment with different chemical reagents showed different colour. Phytochemical evaluation revealed the presence of triterpenoids, saponins, tannins, flavonoids and steroids. Keywords: Cleome viscosa, Dog mustard, Microscopy, Pharmacognosy, Physicochemical. IPC code; Int. cl. ( ) A61K 36/00 Introduction Ethnomedically, the whole plant and its parts (leaves, seeds and root) are widely used by local people for the treatment of various disease conditions without standardization. The standardization of a crude drug is an integral part of establishing its correct identity. Before any crude drug can be included in an herbal pharmacopoeia, pharmacognostic parameters and standards must be established. Microscopic method is one of the simplest and cheapest methods to start with for establishing the correct identity of the source materials 1. Cleome viscosa Linn. (Family: Capparaceae) is a weed distributed throughout the tropics of the world and the plains of India. The plant is an annual, sticky herb with a strong penetrating odour, yellow flowers and long slender pods containing seeds. It is known as Hurhur (Hindi), Hurhuria (Bengali), Nayikkadugu (Tamil) in Indian traditional medicine 2,3. *Correspondent author: Address: University Institute of Pharmacy, Chhatrapati Shahu Ji Maharaj University, Kanpur , Uttar Pradesh, India herbalprakash@yahoo.com; Telephone: Uses in traditional medicine and other reported activities In Ayurvedic system of medicine, the plant is used in fever, inflammations, liver diseases, bronchitis and diarrhoea 4. The rural people use the fresh juice of the crushed seed for infantile convulsions and in mental disorders. The juice of the plant diluted with water is given internally in small quantities in fever and the leaves are useful in healing the wounds and ulcer 2,5,6. C. viscosa is highly effective in a wide spectrum of diseases and reported to possess antidiarrhoeal, analgesic, psycho-pharmacological, antimicrobial properties including in vitro Helicobacter pylori and wound healing activity However, available literature revealed that no detailed pharmacognostic studies have been carried out on plant; hence the present investigation was undertaken. The object of present study is to evaluate various pharmacognostical parameters such as macroscopy, microscopy, physicochemical parameters, fluorescence analysis and phytochemical studies of the plant. Previously isolated constituents A wide variety of chemical constituents have been isolated from various parts of C. viscosa. A novel

528 INDIAN J NAT PROD RESOUR, DECEMBER 2012 umbeliferone derivative, designated as cleosandrin, series of coumarino-lignans (cleomiscosins) from the seeds and a new glycoside eriodictyol-5-rhamnoside

2 528 INDIAN J NAT PROD RESOUR, DECEMBER 2012 umbeliferone derivative, designated as cleosandrin, series of coumarino-lignans (cleomiscosins) from the seeds and a new glycoside eriodictyol-5-rhamnoside have been isolated from the whole plant 13. Materials and Methods Plant material Whole plant of C. viscosa was collected from the rural area around Kanpur, India. The whole plant material was identified, authenticated taxonomically by Dr. Tariq Husain, Taxonomist of National Botanical Research Institute (NBRI), Lucknow, India and a voucher specimen (No. UIOP/M-121) was deposited at the herbarium section of departmental museum for future reference. Pharmacognostic evaluation Fresh whole plant of C. viscosa (Plate 1) was taken for morphological and histological studies. Coarse powder (60 #) was used to study microscopical characters, physicochemical parameters and phytochemical investigation. For the microscopical studies, transverse sections of leaves, petiole, stem and root were prepared and stained as per standard procedure 14,15. The powder microscopy was performed according to the method of Khandelwal 16. Physicochemical and phytochemical analysis Physicochemical values such as percentage of ash values and extractive values were determined according to the well established official method and Plate 1 Cleome viscosa plant procedure 17,18. Preliminary phytochemical screening was carried out using the standard procedure described by Khandelwal 16. Fluorescence analysis Powdered leaf, root and seed material were treated with various chemical reagents and exposed to visible, ultraviolet light (Short UV) to study their fluorescence behavior 19. HPTLC fingerprint profile For proper and meaningful utilization it is important to have quality standards of material and for this quality standardization HPTLC finger print profile of methanolic extract of C. viscosa (10 µl of 1 mg/ml) was developed. The HPTLC analysis was carried out on precoated silica gel G 60 F 254 TLC plate (Merck) with the help of Camag Linomat IV applicator. The plate was eluted with toluene: ethyl acetate: glacial acetic acid (5:5:0.3) as mobile phase. After development, the plate was dried and densitometrically scanned on a TLC scanner III at 366 nm using Wincat software (CAMAG, Switzerland) and peak area was recorded. Results and Discussion Macroscopic characters Macroscopically, the fresh leaf of C. viscosa was green in colour, alternate, palmately compound with 3-5 leaflets, 2-6 cm long, 1-3 cm wide, leaf stalk mm in length with hairs, obovate to lanceolate in shape with entire margin, obtuse to acute apex, rounded to acute base. Flowers bisexual, yellow in colour; fruit cylindrical, erect capsule, many seeded; seeds reddish brown in colour, sub-orbicular with narrow cleft covered with strong cross ribs and faint concentric ribs; stem angular, sticky with hairs, unbranched to sparsely branched; root long, tap with some secondary roots. Plant has characteristic smell, bitter taste and is sticky in nature. Microscopical characteristics Leaf T.S. of leaf (Plate 2) passing through midrib region show slight depression on upper side and broad cushion on lower side. Single layered, thick walled upper and lower epidermis covered with thick cuticle, multicellular glandular and simple covering trichomes (Plate 3a). Meristele consists of collateral vascular bundle; ground tissue collenchymatous. T.S. passing through lamina region show 2 to 3 layer of chlorophyllous palisade cells followed by spongy

of petiole (Plate 4) is pear shaped with deep depression at upper side; single layered epidermal cell covered with thick cuticle; ground tissue collenchymatous embedded with six vascular bundle.

3 GUPTA & RAO: PHARMACOGNOSTICAL STUDIES OF CLEOME VISCOSA LINN. 529 mesophyll. Lateral vascular bundle also observed, embedded with spongy mesophyll and anomocytic type of stomata (Plate 3b) present in both the surfaces of leaf. Petiole T.S. of petiole (Plate 4) is pear shaped with deep depression at upper side; single layered epidermal cell covered with thick cuticle; ground tissue collenchymatous embedded with six vascular bundle. Vascular bundle is collateral and capped with sclerenchymatous bundle sheath. Trichomes present on the surfaces of epidermis; stalk is multicellular with small globular head. Stem T. S. of stem (Plate 5) almost circular with wavy out line and four larger processes. Single layered and thick walled epidermis covered with thick cuticle; multicellular trichomes present on the surfaces; hypodermis collenchymatous followed by 2 to 3 layer of collenchyma; cortex parenchymatous; pericycle present in form of group of sclerenchyma and pericyclic fibres are in arc shape below each notch. Phloem well developed in form of ring and consist of sieve tubes, companion cells and phloem parenchyma; cambium not distinct. Xylem is present in form of continuous ring and consists of vessels, tracheids, fibres and xylem parenchyma; vessels are in radial rows. Medullary rays are distinct; centre portion occupied by collenchymatous pith. Root T. S. of root (Plate 6) almost circular in outline; the outer most layers is thick walled, remnant of cork cells which become crushed at places and cortex Plate 2 T.S. of Cleome viscosa leaf Plate 3 T.S. of leaf showing; a-covering trichome, b-anomocytic stomata

530 INDIAN J NAT PROD RESOUR, DECEMBER 2012 consist of thin walled mostly oval shaped parenchymatous cells; phloem is well developed and consist of sieve tubes, companion cells and phloem parenchyma.

of Cleome viscosa petiole Powder microscopic characters The powdered leaf and stem material was greenish in colour; showing fibres (Plate 7a), pitted vessels (Plate 7b), spiral reticulate vessels

The powdered root was straw in colour; showing fragments of cork cells with phelloderm (Plate 8a), parenchyma (Plate 8b), medullary rays crossing the parenchyma (Plate 8c), and reticulate thickned

4 530 INDIAN J NAT PROD RESOUR, DECEMBER 2012 consist of thin walled mostly oval shaped parenchymatous cells; phloem is well developed and consist of sieve tubes, companion cells and phloem parenchyma. Major portion of the root is occupied by secondary xylem which consists of vessels, tracheids, fibres and xylem parenchyma; uni to biseriate medullary rays are also distinct. Plate 4 T.S. of Cleome viscosa petiole Powder microscopic characters The powdered leaf and stem material was greenish in colour; showing fibres (Plate 7a), pitted vessels (Plate 7b), spiral reticulate vessels (Plate 7c), fragments of spongy mesophyll (Plate 7d), palisade cells, glandular trichome (Plate 7e), epidermal cells with vessels and calcium oxalate prism (Plate 7f). The powdered root was straw in colour; showing fragments of cork cells with phelloderm (Plate 8a), parenchyma (Plate 8b), medullary rays crossing the parenchyma (Plate 8c), and reticulate thickned vessels (Plate 8d). Preliminary phytochemical screening Preliminary phytochemical screening mainly revealed the presence of triterpenoids, saponins, tannins, flavonoids and steroids. Plate 5 T.S. of Cleome viscosa stem Plate 6 T.S. of Cleome viscosa root [Abbrevations used in plates 2-6: Ep: Epidermis; Uep: Upper Epidermis; Lep: Lower Epidermis; Cu: Cuticle; C: Cork; Ct: Cortex Pal: Palisade Cell; Scl: Sclerenchyma; Col: Collenchyma; M: Mesophyll; Ph: Phloem; Xy: Xylem; VB: Vascular Bundle; PF: Pericyclic Fibre; V: Vessels; P: Pith; GT: Glandular trichome; T: Trichome] Physicochemical parameter Ash values of drug give an idea of earthy matter or the inorganic composition and other impurities present along with drug. Extractive values are primarily useful for the determination of exhausted and adulterated drugs 20. Foreign matter, loss on drying, extractive value and ash analysis of powdered leaf, root and seed of C. viscosa were carried out and comparative results are shown in Figure 1. Fluorescence analysis of powdered drug was also observed and the results are shown in Table 1. As there is no detailed pharmacognostic study on record of this drug which is of great value, present study was taken up with a view to lay down the microscopic standards, which could be used in deciding the genuineness of the drug source. The drug was found to contain simple and multicellular glandular trichomes and trichomes are epidermal outgrowths of considerable value for taxonomic purposes. The presence of 5 to 6 collateral vascular bundle caped with sclerenchymatous bundle sheath in petiole and arc shaped pericyclic fibre in stem and distinct uni- to biseriate medullary rays of root are some of the diagnostic features noted from anatomical study of the plant. Ash values and extractive values

GUPTA & RAO: PHARMACOGNOSTICAL STUDIES OF CLEOME VISCOSA LINN.

b-pitted vessels, c-spiral reticulate vessels, d-spongy mesophyll,

characteristics of Cleome viscosa root: a-cork cell with phelloderm,

5 GUPTA & RAO: PHARMACOGNOSTICAL STUDIES OF CLEOME VISCOSA LINN. 531 Plate 7 Powder characteristic of Cleome viscosa leaf: a-fibres, b-pitted vessels, c-spiral reticulate vessels, d-spongy mesophyll, e-glandular trichome, f-calcium oxalate prism Plate 8 Powder characteristics of Cleome viscosa root: a-cork cell with phelloderm, b-parenchyma, c-medullary rays crossing the parenchyma, d-reticulate thickned vessels

6 532 INDIAN J NAT PROD RESOUR, DECEMBER 2012 Fig. 1 Comparative physicochemical studies of different parts of Cleome viscosa Table 1 Fluorescence analysis of leaf, root and seed powder of Cleome viscosa Treatment Observation under day light Observation under UV light (254 nm) Leaf Root Seed Leaf Root Seed Powder (P) as such Green Straw Brown Dark green Pale green Coffee P + 1N HCl Green Straw Auburn Herbage green Jelly green Greenish black P + HNO 3 Fulvous Golden brown Brown Green Fluorescent green Greenish black P + H 2 SO 4 Olive Dark brown Brown Green Blackish green Blackish green P +1N NaOH in Water Green Straw Chocolate Green Jelly green Dark green P +1N NaOH in Methanol Pale green Straw Olive brown Green Fluorescent green Greenish black can be used as reliable aid for detecting adulteration. These studies help in identification and authentication of the plant materials. Percentage extractive and ash analysis were carried out and results showed that total ash of leaf was about two times higher than root and about four times higher than seed. Alcohol soluble extractive value of seed was two times higher than water soluble extractive value and water soluble extractive value of leaf and root was more than two times higher than alcohol soluble extractive value. Extractive values are useful to evaluate the chemical constituents present in the crude drug. The fluorescent analysis under day light and UV light by treatment with different chemical reagents showed different colour. This analysis suggests that, leaves, root and seed extract of C. viscosa probably contain active agent(s) and this provides the basis for their folkloric use as a cure for some human ailments. HPTLC studies The preliminary HPTLC studies revealed that the solvent system toluene: ethyl acetate: glacial acetic acid (5:5:0.3) was ideal for the methanolic extract of C. viscosa and gave well resolved peaks. The band in the sample were obtained at R f 0.26, 0.38, 0.44, 0.49, 0.72 and 0.79 which can be used as identifying marker (Plate 9).

7 GUPTA & RAO: PHARMACOGNOSTICAL STUDIES OF CLEOME VISCOSA LINN. 533 Plate 9 HPTLC profile and densitometric scanning of methanolic extract of Cleome viscosa. Solvent system: toluene: ethyl acetate: glacial acetic acid (5:5:0.3), Detection: Under UV light λ 366 nm Conclusions The present work was undertaken with an aim of pharmacognostic investigation of C. viscosa providing useful information, which could be useful to detect the authenticity of this medicinally useful plant. Pharmacognostic evaluation can be useful to substantiate and authenticate the drug. Acknowledgement Authors thank Dr Sayyada Khatoon, Scientist E1, National Botanical Research Institute, Lucknow, for her kind help in identifying the microscopical features. References 1 Singh S, Machawal L and Chauhan M G, Pharmacognostic study of male leaves of Trichosanthes dioica with special emphasis on microscopic technique, J Pharmacog Phytother, 2010, 2, Nadkarni A K, Indian Materia Medica, Popular Prakashan, Bombay, 1982, Vol. 1, pp , Asolkar L V, Kakkar K K and Chakre O J, Glossary of Indian Medicinal Plants With Active Principles, Publication and Information Directorate, Council of Scientific and Industrial Research, New Delhi, 1992, p Chatterjee A and Pakrashi S C, The Treatise on Indian Medicinal Plants, Council of Scientific and Industrial Research, New Delhi, 1991, 2 nd Edn, Vol. 1, p The Wealth of India-A Dictionary of Indian Raw Material and Industrial Products Raw Materials, Publications and Information Directorate, CSIR, New Delhi, 1950, Vol. II, p Kirtikar K R and Basu B D, Indian Medicinal Plants, Lalit Mohan Basu Publication, Dehra Dun, 1984, 2 nd Edn, Vol. 1, pp Parimala Devi B, Boominathan R and Mandal S C, Evaluation of anti-diarrhoeal activity of Cleome viscosa extract in rats, Phytomedicine, 2002, 9, Mandal S C, Parimala Devi B and Boominathan R, Studies on analgesic activity of Cleome viscosa in mice, Fitoterapia, 2003, 74, Parimala Devi B, Boominathan R and Mandal S C, Studies on psychopharmacological effects of Cleome viscosa Linn. extract in rats and mice, Phytother Res, 2004, 18, Sudhakar M, Rao Ch V, Rao P M and Raju D B, Evaluation of antimicrobial activity of Cleome viscosa and Gmelina asiatica, Fitoterapia, 2006, 77, Mahady G B, Bhamarapravati S, Adeniyi B A, Pendland S L, Doyle B, Locklear T and Solver C, Traditional Thai medicines inhibit Helicobacter pylori in vitro and in vivo: Support for ethnomedical use, Ethnobot Res Appl, 2006, 4, Panduraju T, Parvathi B, Rammohan M and Srinivas Reddy C, Wound healing properties of Cleome viscosa Linn., Hygeia J D Med, 2011, 3, Mali R G, Cleome viscosa (wild mustard): A review on ethnobotany, phytochemistry and pharmacology, Pharm Biol, 2010, 48, Brain K R and Turner T D, The Practical Evaluation of Phytopharmaceuticals, Wright-Scientechnica, Bristol, 1975, pp Kokate C K, Practical Pharmacognosy, Vallabh Prakashan, Delhi, 4 th Edn, 1994, pp Khandelwal K R, Practical Pharmacognosy, Nirali Publication, Pune, 19 th Edn, 2008, pp

8 534 INDIAN J NAT PROD RESOUR, DECEMBER World Health Organisation (WHO), Quality Control Methods or Medicinal Plant Material, Organisation Mondiale De La Sante, Geneva, 1992, p Anonymous, Indian Pharmacopeia, Ministry of Health and Family Welfare, Controller of Publications, New Delhi, 1966, 4 th Edn, A53-A Pratt R J and Chase C R, Fluorescence of powdered vegetable drug with particular reference to development of a system of identification, J Amer Pharm Assoc, 1949, 38, Gupta P C, Sharma N and Rao Ch V, Pharmacognostic studies of the leaves and stem of Careya arborea Roxb., Asian Pac J Trop Biomed, 2012, 2(5),

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