A credit seminar on

Isolation and Characterization of Secondary metabolites

from leaves of Carica papaya Linn.

Advisor

Prof. (Dr.) Inder Pal Singh

Presented by

Gaurav

M.S.(Pharm.) II Semester

Department of natural product (TM)

National Institute of Pharmaceutical Education and Research

S.A.S. Nagar

FLOW OF PRESENTATION

1. INTRODUCTION

2. AIM & OBJECTIVE

2.1. Aim

2.2. Objective

3. LITERATURE REVIEW

4. OUTLINE FOR WORK PLAN

1- Introduction

The spread of knowledge on the natural wealth is more important

for a country like India, at a time when the synthetic drugs are

stealing the economy rates.

The limited knowledge on the varied use of the medicinal plants,

their analytical profile , clinical research data , their availability

and extent of distribution weakens the ways to utilize these

resources efficiently. Therefore, it is required to bring the

information in various sources into one roof.

Due to their natural origin, less side effect and high chemical

constituents these are highly utilizing in developed and developing

countries.

According to the ethnobotanical Carica papaya Linn. used

traditionally in various disease to protect and maintain good health.

Abo.K.A., Fred.J, et al, J. Ethno pharmacology 2007; 115: 67–71

Papaya is a powerhouse of nutrients and is

available throughout the year. Papaya belongs to

the family Carica papaya Linn. It is a rich source

of three powerful antioxidant, vitamin C, vitamin

A and vitamin E. The minerals, magnesium and

potassium, vitamin B pantothenic acid and foliate

and fiber. In addition to all this, it contains a

digestive enzyme -papain that effectively treats

causes of trauma, allergies and sports injuries.

And also used as anti malarial , anti ulcer, anti

hypertensive activity, wound healing activity,

hepato-protective activity etc.

Leaf

T.Vij, Y.Prashar, et al ,A.Pac.J.Trop.Dis. 2015; 5: 1-6

Dulce M Rivera-Pastrana,et al, J.Sci. Food Agric .2010; 90: 2358–2365

https://www.evaidya.com

2.1- Aim – Isolation and characterization of secondary metabolites from

leaves Carica papaya Linn.

2.2- Objectives

Collection & Authentication of plant material

Preparation of aqueous methanolic extract

TLC profiling of prepared extract

Isolation and identification of compounds from aqueous methanolic extract

of Carica papaya Linn. leaves using various chromatographic techniques.

Characterization of isolated compounds using modern spectroscopic

methods.

2. AIM & OBJECTIVE

3.1- Taxonomical classificationKingdom : Plantae

Division : Magnoliophyta

Class : Magnoliopsida

Order : Brassicales

Family : Caricaceae

Genus : Carica

Species : Carica papaya Linn.

3.2- Vernacular namesEnglish : Papaya, pawpaw, papaw,

Hindi : Papita,

Sanskrit : Arand-kharpuja,

Marathi : Pappay

3- LITERATURE REVIEW

T.Vij, Y.Prashar, et al ,A.Pac.J.Trop.Dis. 2015; 5: 1-6Dulce M Rivera-Pastrana,et al, J.Sci. Food Agric .2010; 90: 2358–2365

3.3- Description

Height : 10 to 15 feet

Spread : 5 to 7 feet

Plant habit : Upright

Growth rate : Fast

3.3.1- Leaf

Leaf color : Green

Leaf arrangement : Alternate

Leaf type : Simple

Leaf shape : Star-shaped

Leaf venation : Palmate

Leaf blade length : 18 to 36 inches

3.3.2- Flower

Flower color : White- yellow

Flower characteristic: winter flowering,

spring flowering.

3.3.3- Fruit

Fruit shape : Oval

Fruit length : 6 to 12 inches

Fruit color : orange; yellow

T.Vij, Y.Prashar, et al ,A.Pac.J.Trop.Dis. 2015; 5: 1-6

Dulce M Rivera-Pastrana,et al, J.Sci. Food Agric .2010; 90: 2358–2365

https://www.evaidya.com

Geographical Distribution

Carica papaya Linn. Cultivation and

collection include Hawaii, Tropical

Africa, the Philippines, India,

Ceylon, Malaysia and Australia,

apart from the widespread but

smaller scale production in South

Africa, and Latin America. In India,

papaya is cultivated in Maharashtra,

Bengal, Bihar, Haryana, Punjab,

Delhi, Andhra Pradesh and Uttar

Pradesh. Cultivators of Carica

papaya Linn are described. These

were found in different varieties

with different characteristic.

http://www.mapsofindia.com/indiaagriculture/frui

ts-map/papaya-producing-states.html

V. Yogiraj, P. K.Goyal, et al, I.J.H. Med 2014; 2 : 01-08

Phytochemical constituents present in Carica papaya Linn.

In phytochemical study of Carica papaya Linn. which mainly include different

type of the chemical compounds .These compounds include alkaloids,

flavonoids, Quinons, Saponin glycoside, Enzymes, Amino acids, Phenols and

phenolic acid, Organic acids, and others like creatine, cholin,Trigonelline etc.

FIGURE No.- (1) PAPAIN (2) LYCOPENE.

M.Azarkan ,et al,“J.chrom. B, 2003;790: 229–238

N. Otsuki, N.H. Dang, et al, J. Ethnopharmacol. 2010;127: 760–767.

FIGURE No.-(3) CARPAINE (4) DEHYDROCARPAINE I (5) DEHYDROCARPAINE II

(6) EMETINE

Chung-Shih Tang, et al, Phytochemistry, 1979; 18: 651-652

Julianti T, et al. Planta Med 2014; 80: 1138-42K. Kovendan, K. Murugan, et al, A.Pac.J.Trop.Dis.(2012)S306-S311

V.Zunjar, et al, J.Ethnopharmacol. 2015(http://dx.doi.org/10.1016/j.jep.2016.01.035)

The quantity of carpaine in water, petroleum ether and ethyl acetate extract was found to be 16.79 ± 0.09 mg/g, 117.10 ± 0.59 mg/g, 99.47 ± 0.53 mg/g.and Qty of other(3)0.15%(4)0.33% (5)0.51% (6) 0.15- 30%

FIGURE No.- (7) Quercetin 3-(2G-rhamnosylrutinoside) (8) Kaempferol 3-(2G-rhamnosylrutinoside) (9)

Quercetin 3-rutinoside (10) Myricetin 3-rhamnoside

A.Nugroho, et al, A. Pac. J.Trop. Biomed 2017; 7: 208–213.

Chen.H., Zuo.Y., Deng.Y, et al, J.Chrom. A 2013: 913, 387–395Rivera-Pastrana.D.M., et al, J. Sci. F. Agric, 2010; 90: 2358–2365.

Content of compounds in MeOH extract of C. papaya leaf mg/g(7)3.11 (8) 7.23 (9) 0.97 (10) 0.81

Figure No. (11) Caffic Acid (12) Ferulic acid (13) 5,7 Dimethoxycoumarin (14) Protocatechuic acid .

A.Caninia, D.Alesiania,et al, J. F. Comp.Analysis 2007;20: 584–590Chen, H., et al , J.Chrom. A 2001; 913, 387–395N.Gonga ,et al, J.Pharm.Biomed.analysis 2015; 115: 74-85Dulce M Rivera-Pastrana,et al, J.Sci. Food Agric .2010; 90: 2358–2365

Qty of compounds in mg/g(11) 0.25(12) 0.19(13) 0.33(14) 0.11

Figure No. (15) p-Coumeric acid R1=R3=R4=H,R2=OH (16) O-Coumaric acid R1=R2=R3=H,R4=OH (17) Chlorogenic acid (18) Quercetin (19) Kaempferol.

A.Nugroho, et al, A. Pac. J.Trop. Biomed 2017; 7: 208–213

A.Caninia, D.Alesiania,et al, J. F. Comp.Analysis 2007;20: 584–590Chen, H., et al , J.Chrom. A 2001; 913, 387–395N.Gonga ,et al, J.Pharm.Biomed.analysis 2015; 115:74-85

The Qty of compounds-mg/g in leaf(15) 0.33(16) 0.29(17) Trace qty(18) 0.03(19) 0.04

3.6- Pharmacological properties of C. papaya Linn

The plant C. papaya has been proved for various medicinal activitieslike antioxidant, anti-hypertensive, wound healing, hepatoprotective,anti-inflammatory, antimicrobial, antifungal, anti-fertility,histaminergic, diuretic, anti-amoebic, anti-tumor, anthelmintic, effecton smooth muscles, antimalarial, hypoglycemic activity, immuno-modulatory activity, anti-ulcer activity, anti -sickling activity.

3.6.1- Antioxidant activityThe methanolic extract of unripe fruits of C. papaya was evaluated invivo for its effect on activities of some antioxidant enzymes whichincludes glutathione peroxidase(GPx), glutathione transferase (GST),glutathione reductase, catalase and glucose-6-phosphatedehydrogenase in mice treated with a orally dose of 100 mg/kg.potential.11,14

A.Nugroho, H.Heryani, et al, A.P.J.Trop. Biomed, 2017; 7:208-213J.N.Calvache.et al, J.F.Foods, 2016; 27: 319-328Ze-You Li, Y.Wang,et al, A.P. J.Trop. Med., 2012; 5: 231-233N.Otsuki,et al, J.Ethnopharmacology, 2010; 127: 760-767

3.6.2- Anti-hypertensive activityThe ethanolic extract of unripe fruit of C. papaya (20 mg/kg, i.v) wasused for the anti-hypertensive which produced a significantdepression of MAP in normotensive, renal and DOCA-salthypertensive animals groups as scompared to control.

3.6.3- Wound healing activityThe aqueous extract of C. papaya fruit [100 mg/kg. for 10 d] forwound healing. The aqueous extract shows 77% reduction in thewound area when compared to 59% contraction to wound of thecontrols.

3.6.4- Hepatoprotective activityThe aqueous (250 mg/kg, p.o) and ethanol (250 mg/kg, p.o) extractsof C. papaya showed significant hepatoprotection by lowering thebiochemical parameters such as SGPT, SGOT, serum bilirubin,akaline phosphatase.

Shila Gurung,et al, J.Ethnopharmacology 2009; 121: 338–341Sripanidkulchai.B,et al, J Ethnopharmacol. 2001;75: 185-90.

3.6.6- Antimicrobial activity11

The aqueous extract of C. papaya leaves and roots at differentconcentrations (25, 50, 100, 200 mg/mL) showed antimicrobial activityagainst some human pathogenic bacteria using the agar diffusionmethod.

3.6.7- Antifungal activityThe latex of C. papaya and fluconazole has synergistic action on theinhibition of Candida albicans growth. This synergistic effect resultsin partial cell wall degradation. Latex proteins appear to beresponsible for antifungal action and minimum protein concentrationfor producing a complete inhibition was reported as about 138mg/mL.

T. Kokila, P.S. Ramesh,et al,J.Ecotoxicol.Environ.Saf. 2016; 134: 467-473R.Chandrasekaran, S.Gnanasekar.et al, J.Mol.Liq., 2016; 219 232-238Osato.J.A., Santiago.L.A., Remo.G.M.et al, Life Science 53, 1383–1389.

3.6.8- Anti-fertility activityThe crude extract of bark of C. papaya [5-10 mL/(kg. d), p.o for 4 weeks] on the seminiferous tubules of rats showed complete loss of fertility attributing to decline in sperm motility and alteration in their morphology.

3.6.9-Anti- Histaminergic activityThe crude extract of C. papaya (0.5-512 µg/mL) show anti histaminergic activity by acting on H1 receptor.

3.6.10- Diuretic activityAqueous root extract of C. papaya when given orally at the dose of 10 mg/kg to rats produced significant diuretic activity.

3.6.11- Anti-malarial activityThe highest larval mortality in the ethanol leaf extract of C. papay against P. falciparum at different dose concentrations (25, 50,100 and 150 μg/mL) exhibited promising inhibitory activity .

E.Julaeha, Y.Permatasari, et al, J.Procedia Chem. 2015; 17: 66-69E.Hainida, K.Ikram, J.Food Composition and Analysis 2015; 41: 201–211T.Vij, Y.Prashar, et al, A.Pac. J.Trop.Dis. 2015; 5: 01-06K. Kovendan, K. Murugan, C. Panneerselvam, et al, A.Pac.J.Trop.Dis. 2012; 2: 306-311

3.6.12- Hypoglycemic activityThe ethanolic leaf extract of C. papaya at the dose 5.0 mg/ kgproduced significant blood sugar level reduction with no significanteffects at the higher dose of 10 mg/kg. Increased the hypoglycemiceffect of metformin with the variables interacting differently for eachdrug-extract combinations.

3.6.13- Anti-ulcer activityAqueous seed extract of C. papaya at the dose of 50 mg/ kg and 100mg/kg p.o against alcohol induced acute gastric damage and bloodoxidative stress in rats. The gastric acidity was significantly reducedin rats treated with 100 mg/ kg of the extract.

Hussein O.B. Oloyede, Matthew C. Adaja,et al, J.Integr. Med., 2015; 13: 105-114I.E.Juárez-Rojopa,et al, Rev Bras Farmacogn 2014; 24: 341-347 Abo.K.A., Fred.J, et al, J. Ethno pharmacology 2007; 115: 67–71

1

•1:Collection and authentication of plant material.

•Drying and powdering

2

•Preparation of aqueous methanolic extract.

•Fractionation.

3

• Isolation of compound using suitable stationary and mobile phase by chromatography method (CC, FLC, VLC etc ).

•Purification of compound.

4

•Structure Elucidation after isolation and purification by spectroscopy methods (UV, NMR , IR etc.)

•Submission of compound .

OUT LINE FOR WORK PLANE-