Annona Squamosa)

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Jaswinder Kaur, Amit Mittal, Saurabh Singh, Dileep Singh Baghel* School of Pharmaceutical Sciences, Lovely Professional University, Jalandhar - Delhi G.T. Road, Phagwara, Punjab (India)-144411

ABSTRACT Herbal remedies were utilized from ancient time for the treatment and prosperity of human beings. A portion of the regular therapeutic plants are common to the point that we utilized them in every day existence without knowing their restorative significance. Sitaphal is one among of them usually known as custard apple, sugar apple; sweetsop is the fruit of Annona squamosa (Annonaceae). The leaves of the plant have been utilized as bug spray, anthelmintic and in healing of bleeding wounds. Fruit of custard apple is a tropical fruit mainly grow in tropical climate. The fruits are well known for their delicious taste and are heart shaped with light green skin, a soft creamy white flesh. Nutritionally, the fruits are rich in carbohydrates mainly in the form of sugar (23.5%), protein (1.6%), calcium (70mg/100gm), phosphorous (47mg/100gm) and iron (1.5mg/100gm). It is said to contain the qualities of the rejuvenating drugs. It is found to be very useful for brain and nervous system. It also enhances muscular strength and tone up the heart. Annona species have been widely grown throughout Central and South America. It is also grown in West Indies, Taiwan, USA other than India. It contains various chemical compounds such as alkaloid, isomeric hydroxyl ketones etc. and squamone from bark of it which show antibacterial, antidiabetic, antitumor, anti-malarial, anthelmintic, hepatoprotective, antioxidant, respiratory stimulant and have diuretics properties. KEYWORDS: Annona, custard apple, phytoconstituents, sitaphal INTRODUCTION The name sitaphal is originated [1] from the two Sanskrit words “sita” means cold and “phal” means fruit. It belongs to family annonaceae. It is regarded by way of numerous synonyms names including sugar apple, sweet cleaning soap, sitaphal and sharifa. Custard apple prospers properly in tropical and warmer sub tropics elements of India. It is cultivated in Maharashtra, Punjab, Bihar, Assam. It grows better in the dry region and in heavy rainfall. Fruit of sitaphal is rich in nutrition. Various pharmacological studies done on this plant. The plant grows better in the month of April [2]. It contains various chemical constituents. This plant is popular in a Ramayana period. Fruit of sitaphal is a rich source of sugar. Sitaphal also have an anti-ageing property. It provide glow to the skin [3]. Fruiting process starts after 2-3 years of germination. It grows better in the sandy soil. The leaf extract of sitaphal also reported to contain anti-bacterial properties against gram positive and gram negative bacteria [4]. During Ramayana period Mata Sita eat this fruit so this plant is named as sitaphal [5].

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) Parts of plant Morphological characters Leaves Deciduous, rectangular, slender-lanceolate Fruit Compound, heart shaped, abnormal, yellow or brown color when ripped Seed Dark brown or black in color, glossy, oblong and smooth Flower Flowers present drooping clusters, fragrant, slender, greenish yellow in color SYNONYMS [6] English : custard apple,sugar apple,sweet sop Sanskrit : seetaphalam,sudha,subha Hindi : seetaphal,sharifa,ata Malayalam : athachakka,atha,seetapazham Kanda : sitaphal Telgu : sitaapandu Tamil : sitapplam Bengali : ata Arabic : ghista Italian : pomo canella French : pomme de cannelle,attire Portuguese : atta,fructa do conde TAXONOMIC CLASSIFICATION [7, 8] Annona squamosa L. Kingdom : Plantae Subkingdom :Tracheobionta Super division : Spermatophyta Division : Magnoliophyta Class : Magnoliopsida Sub class : Magnoliidae Order : Magnoliales Family : Annonaceae Genus : Annona L. Species : Annona squamosa

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) AYURVEDIC PROPERTIES [6] Rasa: madhur,kashaya Guna: guru,snigdha Virya: sheet Vipaka: madhur Karma: udar rog, hridya etc. HISTORY In the mythological literature of Hinduism, it is mentioned that the lord Rama who killed Ravana due to abducting lord Rama’s wife Sita mata, he kept mata sita at ashoka vatika at lanka which is now situated in Srilanka. In ashoka vatika there were many different varieties of fruits and plants were present, Mata Sita used to have those fruits as a meal one of the main fruits was sitaphal. This fruit got renowned because of Sita mata eat it so named as sitaphal [1]. In few literatures it is mentioned that at the time of abduction of Sita mata by Ramavana, Sita mata was crying and her tears fall down on the ground, which result in production of sitaphal trees. Due to the tears of Mata Sita this fruit named as sitaphal [1].

CHEMICAL CONSTITUENTS [9-13] Fruits: Sugar (up to 28%), Vitamin C, Iron, Calcium, Thiamine, Amino acid, Potassium, Carotene, Riboflavin, Niacin, Ascorbic acid, Magnesium and dietary fibers. Leaves: Isoquinoline alkaloids, Acetogenine (annoreticulin and isoannoreticuin), Dopamine alkaloid, Salsolnol, Coclaurine, Carvone, Eugenol, Farnesol, Geraniol, Limonene, Linalool, Menthone, Atrophine, Roemerine, norisocoryline and sugars like rhamnoside and quercetin-3-glycoside. Leaf oil yielded 59 chemical compounds. Roots and Stem: Liriodenine, Oxoanalobine, Borneol, Camphene, Camphor, Car-3-ene, β-Caryphyllene, Eugenol, Farnesol, Geraniol, 16-Hetriacontanone, Hexacontanol, Higemamine, Isocorydine, Limonine. Bark: Bullatacin, N-Nitrosoxylopine, Roemerolidine, Duguevalline, Mosinone-A, Mosin-B, Mosine-C, Squamone. Seeds: Acetogenin, Solamin, triterpenoids (stigmasterol and sitosterol), Annotemoyin-1, Annotemoyin-2, Squamocin, cholesteryl, glucopyranoside.

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Principle Nutrient Value Percentage of RDA

Energy 101 Kcal 5%

Carbohydrates 25.20 g 19%

Protein 1.70 g 3%

Total Fat 0.60 g 3%

Cholesterol 0 mg 0%

Dietary Fiber 2.4 g 6%

Vitamins

Niacin 0.500 mg 3.5%

Pantothenic acid 0.135 mg 2.5%

Pyridoxine 0.221 mg 17%

Riboflavin 0.100 mg 8%

Thiamin 0.80 mg 7%

Vitamin A 33 IU 1%

Vitamin C 19.2 mg 32%

Electrolytes

Sodium 3 mg <1%

Potassium 382 mg 8%

Minerals

Calcium 30 mg 3%

Iron 0.71 mg 9%

Magnesium 18 mg 4.5%

Manganese 0.093 mg 4%

Phosphorus 21 mg 3%

Phyto-nutrients

Epicatechin 5.6 mg --

Proanthocyanidin monomers 6.2 mg --

Proanthocyanidin dimers 14.2 mg --

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PHARMACOLOGICAL PROPERTIES OF ANNONA SQUAMOSA

1. Anti-arthritic, Anti-inflammatory and Analgesic Activity: Singh S screened and reported that the plant extract shows dose dependent analgesic and anti-inflammatory activity but significant reduction in neutrophils infiltration, pannus formation and bone of the animal treated with plant extract [15]. 2. Antioxidant and Antibacterial Activity: El-Chaghaby et al. studied about the effect of different solvent extracts on the antioxidant and antibacterial activities of leaves. Kothari et al. studied about antioxidant activity of seed extracts. Nandhakumar et al. evaluated In l. vitro antioxidant potential of methanol and aqueous extract of fruit pulp. Shenoy et al. reported the antibacterial and wound healing activity of the leaves. [16-18]. 3. Antimalarial Activity and cytotoxic effect: Rahman et al. reported the antimicrobial and cytotoxic constituents present in the seeds of sitaphal. He also reported that the extract of Annona species are potential anti-mosquito agents. In the recent studies on Annona squamosa all compounds showed moderate activity against a chloroquine sensitive strain and a chloroquine resistant strain of Plasmodium falciparum [19]. 4. Anti-ulcer activity : Yadav et al. Identified and investigated the anti-ulcer constituents of Annona squamosa twigs [20]. 5. Anti-tumour activity: Rakesh Ranjan and Mahendra Sahai reported the anti-tumor activities against human hepatoma cells in-vitro and in-vivo, indicating a potential for developing the extract a novel anticancer liver drug. Aqueous extracts of the seeds possess significant anti-tumor activity in-vivo against AD-5 tumor [21]. 6. Hepatoprotective activity: Saleem et al. investigated the hepatoprotective potential of aqueous and alcoholic extract of leaves on experimental animal model 7. Anti-HIV : Saleem et al. reported that the 16β, 17-dihydroxy-ent- kauran-19-oic acid showed significant

activity against HIV replication in H9 lymphocyte cells with an EC50 value of 0.8 µg/ml [22]. 8. Larvicidal activity against Aedes aegypti mosquitoes: R. Kaushik & P. Saini studied about larvicidal activity of acetone leaves extracts. Parvin et al. reported that the chloroform extract of the seeds of have pesticidal activity against both adults and different instars of Tribolium castaneum (Herbst) under laboratory condition. [23, 24]. 9. Anthelmintic activity and Anti-genotoxic Effect: Gupta et a. studied about the anthelmintic activities of the Annona squamosa and its leaf extract have been studied using various models. The hexane, ethyl acetate, ethanolic extracts of the crude drug at different concentrations were tested which involve determination of paralysis time and death time. Antigenotoxic effects of aqueous and ethanolic bark extracts of Annona squamosa was assessed by determining the frequency of micronucleated polychromatic erythrocytes (MnPCEs) and chromosomal aberrations [25].

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) CLIMATE CONDITION [26-28] The plant of sitaphal is well grown in the hot and dry region. Summer rainfall and warm protected soil is most suitable for the growth of the plant. The cold weather and the -fog is not suitable for the growth of sitaphal plant. The plant bears fruits in 3-4 years.

QUALITATIVE TEST FOR PHYTOCHEMICALS [2, 29] Observations Observations Observations Sr. Phytochemical Qualitative Test (Aqueous (Ethanolic (Chloroform No. constituents extract) extract) extract) Ferric chloride test Positive Negative Positive 1 Lead acetate test Tannins Positive Negative Positive Bromine water Positive Negative Positive Borntrager’s test Negative Negative Negative Liebermann’s Test Negative Negative Negative 2 Glycoside Keller-Kiliani Test Negative Negative Negative Salkowski’s Test Negative Negative Negative 3 Foam test Saponin Positive Positive Positive Benedict's solution test Positive Positive Positive 5 Fehling’s test Carbohydrates Positive Positive Positive Molisch’s test Positive Positive Positive Mayer’s reagent Positive Negative Negative 6 Dragendroff reagent Alkaloids Positive Negative Negative Wagner’s reagent Positive Negative Negative

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) Hager’s reagent Positive Negative Negative Liebermann Positive Negative Negative 7 Burchard’s reaction Steroids Salkowski test Positive Negative Negative Ferric Chloride Test Positive Positive Positive Liebermann's nitroso Positive Positive Positive 8 reaction Phenols Lead Acetate test Positive Positive Positive Gelatin test Positive Positive Positive Alkaline reagent test Positive Negative Negative 9 Flavonoids Zinc hydrochloride test Positive Negative Negative TEST DONE ON LEAVES: Chlorophyll test: Chlorophyll a is the photosynthetic pigment that reasons the inexperienced colour in algae and plant life. The awareness of chlorophyll a gift inside the water is directly related to the quantity of algae residing within the water [2]. Instrument used: SPAD SPAD 502 Plus Chlorophyll Meter immediately measures chlorophyll content or “greenness” of your vegetation to reduce the threat of yield-limiting deficiencies or high priced overfertilizing. The SPAD 502 Plus quantifies diffused modifications or tendencies in plant health lengthy earlier than they may be seen to the human eye. Procedure: Taken leaf from a plant, attached that leaf with the chlorophyll meter and measure the range of chlorophyll present in leaf.

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Area of leaves: Sr.no. Leaf area Chlorophyll content 1 27.5 48.8 2 26.0 41.3 3 21.7 40.2 4 17.6 39.5 5 17.5 34.1 This results show that the biggest size of the leaf contain the high chlorophyll content and the smallest one contain the low chlorophyll content. FTIR [30] Significance: 1) It can identify unknown materials. 2) It can determine the quality or consistency of a sample. 3) It can determine the amount of components in a mixture.

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Readings of FTIR: Sr.no. Peak Intensity Area Compound 1 530.44 86.902 1.153 organic halogen compounds (C–X stretching (X = F, Cl, Br or I), phosphorus compounds (P=S stretching, P–Cl stretching)

2 1037.74 88.466 10.886 infrared bands of aliphatic hydrocarbons, infrared bands of aromatic compounds (In-plane C–H bending), Alcohols and Phenols (C–O stretching, O–H) Ethers (C–O), stretching oxygen-containing compounds (C–O stretching) 3 1244.13 93.717 3.051 aromatic compounds (In-plane C–H bending) Alcohols and Phenols, Ethers, oxygen-containing compounds (C–O stretching), Aliphatic (C–O stretching), Anhydrides

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) 4 1379.15 93.932 3.653 nitrogen-containing compounds, Aliphatic nitro compound (NO2 symmetric stretching), Heterocyclic Compounds, Boron Compounds (B–O stretching), 5 1604.83 92.67 5.565 Aliphatic, hydrocarbons, Alkenes 6 1942.38 98.733 0.207 No compound is found at this peak 7 2195.07 98.569 0.198 aliphatic hydrocarbons (Alkynes), organic compounds Combination (N–H stretching), combination (O–H stretching) 8 2920.32 96.095 1.781 No compound is found at this peak 9 3275.24 95.371 0.549 No compound is found at this peak

QUANTITATIVE ESTIMATION OF PHENOL Chemicals required: 1) Folin ciocalteau reagent. 2) 20% sodium carbonate(0.2gm of sodium carbonate in 100ml distilled water. Sample preparation: 1) Take the leaves. 2) Clean them properly. 3) Dry the leaves with the help of tray drier. 4) After drying crush them properly to make fine powder with the help of mortar pestle. 5) Take the powdered sample in test tube. 6) Add 10ml of 80% ethanol in it. 7) Boil it for 10 minutes with the help of water bath. 8) Filter it with the help of filter paper. Procedure: 1) Take 0.25ml of extracted sample in test tube. 2) Boil and evaporate the supernatant extract. 3) Add 5ml of distilled water in above test tube. 4) From the above test tube take 0.2 ml. 5) With the help of distilled water make up the volume upto 3ml. 6) Add 0.5ml folin- ciocalteau reagent. 7) Leave for 3 minutes. 8) Add 2ml of 20% sodium hydroxide. 9) Boil it for 1 minute. 10) Check absorbance at 650nm. Blank sample: JETIRDZ06138 Journal of Emerging Technologies and Innovative Research (JETIR) www.jetir.org 1067

© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) Distilled water. Calculation: (O.D./0.002) mg(100gm) Result: ( 0.147/0.002) =73.5mg(100gm) IN-VITRO ANTIOXIDANT ACTIVITY Chemicals required: DPPH: 0.0236gm of DPPH in 1000ml methanol. Sample preparation: 1) Take the leaves. 2) Clean them properly. 3) Dry the leaves with the help of tray drier. 4) After drying crush them properly to make fine powder with the help of mortar pestle. 5) Take the powdered sample in test tube. 6) Add 10ml of 80% ethanol in it. 7) Boil it for 10 minutes with the help of water bath. 8) Filter it with the help of filter paper. Procedure: 1) Take 0.1ml of extracted sample in a test tube. 2) Add 3.9ml of the DPPH in the test tube. 3) Keep it in dark for 30 minutes. 4) Check the O.D at 517nm. Blank sample: Distilled water and DPPH. Calculation: DPPH O.D. – sample O.D. *100 DPPH O.D. Result: 0.416-0.212/0.416*100 = 50.54 SUMMARY AND CONCLUSION Annona squamosa most famous plant semi deciduous found in the form of shrub it is not very long in height and consists spreading branches. that are usually cultivated all over the World mainly in tropical regions. This plant is having very sweet and edible fruit that is why it is known as custard apple. This plant is not only useful for the edible purposes but also in the field of medicines. According to experiments performed it has been proven that this plant mainly consists chemical constituent’s tannins, phenols, sugars, flavonoids, etc. It has been also observed that this plant possess antioxidant, antibacterial, antimalarial, anti-inflammatory, antiaging and many more. According to literature this plant is also useful in curing dysentery, diarrhoea, rheumatism, colds, chills, and insomnia. Evidence also shows that this plant is having anticancer property and it also

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) suppresses the level of excess uric acid which is present in blood. Plant seeds are having oil in it which produces pesticide activity and extensively used in the field of agriculture. However, this plant having poison in various parts like roots seeds and leaves due to the presence of alkaloids and hydrocyanic acid. However further studies required to prepare effective formulations which can give its contribution in health care system.

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© 2018 JETIR December 2018, Volume 5, Issue 12 www.jetir.org (ISSN-2349-5162) 24. Parvin, M.S., et al., Pesticidal activity of pure compound Annotemoyin-1 isolated from chloroform extract of the plant Annona squamosa Linn. against Tribolium castaneum (Herbst)(Coleoptera: Tenebrionidae). Pakistan journal of biological sciences, 2003. 6(12): p. 1088-1091. 25. Gupta, R.K., et al., Hypoglycemic and antidiabetic effect of ethanolic extract of leaves of Annona squamosa L. in experimental animals. Journal of ethnopharmacology, 2005. 99(1): p. 75-81. 26. Sravanthi, T., Studies on preservation and processing of custard apple (Annona squamosa L.) pulp. ANGRAU ACH: POST GRADUATE & RESEARCH CENTRE. 2004. 27. Hashmi, S.I. and V. Pawar, Studies on physical and chemical characteristics of custard apple fruit pulp from different locations. Journal of Dairying Foods & Home Sciences, 2012. 31(2): p. 117-120. 28. Bhatnagar, P., et al., Evaluation of landraces of custard apple (Annona squamosa L.). Plant Archives, 2012. 12(2): p. 1045-1048. 29. Lohar, D., Protocol for testing Ayurvedic, Siddha and Unani medicines, Government of India Department of AYUSH. Ghaziabad: Ministry of Health & Family Welfare, Pharmacopoeial Laboratory for Indian Medicines, 2007: p. 21. 30. Pavia, D.L., et al., Introduction to spectroscopy. Cengage Learning. 2014.

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