ORIGINAL ARTICLE Proximal and Phytochemical Analysis of Wild Jack Seeds ( hirsutus Lam.), Anti- Diabetic and Anti-Microbial Properties and Formulation of Food Products Krishnapriya K, Kanjana K * Department of Clinical Nutrition and Dietetics, PSG College of Arts & Science, Civil Aerodrome post, Peelamedu, Coimbatore-641014, , .

*Correspondence: [email protected]

ABSTRACT BACKGROUND: The present study aims at evaluating the proximal principles, phytochemicals, anti- diabetic and anti-microbial activity of Lam. (Wild ) seeds thereafter incorporating the seed powder in food. Negligible studies have been conducted to identify the health benefits of A. hirsutus seeds. Hence, an attempt has been made to explore the active principles in the A. hirsutes seeds. METHODS: The seed powder was extracted using three different solvent extracts viz., methanol, di-ethyl ether and water. The extracts were analysed for phytochemicals (qualitative), proximate principles, anti-bacterial (Salmonella typhi, E.coli, Staphylococcus aureus, Klebseilla pneumonia) and anti-fungal (Trichoderma viridae, Aspergillus niger) properties using agar well diffusion method and anti-diabetic properties using α-amylase and α-glucosidase inhibitor activity. Food formulation was done by incorporating the seed powder in traditional foods followed by organoleptic evaluation. RESULTS: The study revealed the presence of phytochemicals such as alkaloids, tannins, sterols and quinine in different solvents and contains greater amounts of carbohydrates and proteins. The study shows the highest anti-microbial activity against Staphylococcus aureus and Trichoderma viridae. The seeds contain potent anti-diabetic activity. CONCLUSION: The study concludes that Artocarpus hirsutus seeds are nutritionally enriched and prevents food borne pathogens. The seeds reveal excellent anti-diabetic properties. The seeds are not consumed frequently due to the lack of information on its health benefits. Therefore, the seeds can be incorporated in traditional foods to improve the nutritional quality and enhance the importance of traditional medicines and foods.

KEY WORDS: Artocarpus hirsutus, Phytochemical, Anti-microbial, Anti-diabetic, Traditional

1. INTRODUCTION Artocarpus is a of approximately of producing a milky sap.Artocarpus hirsutus is 60 and of South-east Asian and a widely acknowledged as a rich source of Pacific origin, belonging to the bioactive secondary metabolites such as family. The name Artocarpus is derived from flavonoids, stilbenes, triterpenoids, and the Greek word artos meaning “bread” xanthones. The ripe fruit of Artocarpus hirsutus andkarpos meaning “fruit”. All Artocarpus is eaten after removing the spiny outer skin. are laticiferous trees or shrubs that are The structure of the fruit is similar to that of composed of , twigs and stems capable the much larger jackfruit. The seeds are also

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edible, usually fried as a snack. Its bark cures medium of cooking using standard procedures diabetes, tapeworm infection, anaemia, suggested by AOAC (2005). The water extract malarial fever, asthma, dermatitis, diarrhoea, of the seed powder was quantitatively pimples and ulcers. It has a long history of analysed for carbohydrate, protein, fibre, medicinal use in Chinese medicine, as a calcium, sodium and potassium. remedy for many kinds of diseases. Artocarpus hirsutus have added a great deal in the field of 2.3 Anti-microbial activity phytochemistry with regard to their availability Anti-bacterial activity of Artocarpus of complex phytochemical components, anti- hirsutus seed extracts were tested against the bacterial activity, anti-helminthic, anti- following bacterial strains: Salmonella typhi, E. inflammatory, and anti-viral[1]. Phytochemical coli, Staphylococcus aureus and Klebsiella screening has been done using the extracts of pnuemoniae. Fungal strains of Aspergillus niger Artocarpus hirsutus , bark and fruit. and Trichoderma viridae were used for anti- Negligible number of studies has been done to fungal screening of the seed extracts. All the identify the health benefits of A. hirsutus seeds. stock cultures were collected from the Centre Hence, an attempt has been made to identify for Bioscience and Nano science Research and explore the active principles present in the Centre, Coimbatore. 24 hr. old pure cultures A. hirsutus seeds. Analysis of proximate active were prepared for use each time. principles that is used in development of food products should be brought to the limelight of 2.4 Bioassay for bacterial strains nutritionists who are on the pathway of The well-diffusion assay was employed improving the community health. in this study. 50 µL of bacterial and fungal cultures aged 18-24 hrs were used. After 2. MATERIALS AND METHODS sterilising the hands with ethanol, the sterilised The wild jack fruit seeds were nutrient agar is poured into 6 petri-plates (4 identified and collected from farmers in the for bacterial screening and 2 for fungal district of Thiruvananthapuram, . The screening). The plates are allowed to solidify. seeds were separated from the fruit and shade The solidified agar medium plates are first dried. Dried seeds were pulverised into fine marked (organism name) under the plate. The powdered form and an approximate amount organism is then spread in each plate was used to prepare extracts with three uniformly using a cotton swab. After this, holes different analytical grade solvents of varying were made using 5 mm cork borer and each polarity such as methanol, di-ethyl ether and hole was filled with 25 µL of extracts. The water. inoculated agar plates were incubated at 37ºC for 24 hrs. the zones of inhibition were then 2.1 Phytochemical Screening recorded in diameter. Two discs were used as The phytochemical analysis was standard, Cefotaxime 30 SDO40 – 1CT performed using all three extracts[1]. Each (bacterial screening) and Ampicillin (fungal extract was subjected to the assays to identify screening). the presence of Alkaloids, Terpenoids, Tannins, Sterols and Quinine. 2.5 Anti-diabetic screening Anti-diabetic activity of the extract was 2.2 Nutrient Analysis tested for α-amylase inhibitory and α- The water extract of the seeds was glucosidase inhibitory activity[6]. analysed for the nutrients as water is the

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2.6 Food formulation 3.2 Anti-microbial Screening In order to formulate food products, In the present study, agar well- the Artocarpus hirsutus seeds were roasted to diffusion method was performed for both anti- remove the outer coat which gives a bitter bacterial and anti-fungal screening. With this taste. After cooling the seeds after roasting, it method, the zone of inhibition for E.coli was was finely powdered. In food formulation, food seen to be highest in aqueous extract with a products were made by combining the seed value of 9 mm (Standard anti-biotic disc – 11 powder in specific proportions in a mm), Staphylococcus aureus in methanol formulation. Incorporation of the seeds extract with a value of 15 mm (Standard anti- powder into South Indian traditional food biotic disc – 7 mm), Klebsiella pneumoniae in products like Idly, Chapatti and Steamed cakes aqueous extract with a value of 10 mm was performed. Three categories of seed (Standard anti-biotic disc – 14 mm) and powder and flour mixtures were formulated. Salmonella typhi in aqueous extract with a The three categories included three different value of 14 mm (Standard anti-biotic disc – 7 proportions of seed powder and flour in the mm). For anti-fungal screening, the zone of ratio of 10:90 (category A), 20:80 (category B) inhibition for Trichoderma viridae in methanol and 30:70 respectively (category C). extract was seen to be highest with a The food products developed were measurement of 8 mm. The anti-bacterial subjected to organoleptic evaluation using 9- screening results are summarized in Table 4. point hedonic scale covering the aspects of Figure 1 shows the different plates and their appearance, texture, flavour, aroma, taste and zone of inhibition. overall acceptability. 3.3 Anti-diabetic Screening 2.7 Analysis of data The results of the anti-diabetic The data collected were processed and screening using both the assays showed very analysed statistically. Sensory evaluation good anti-diabetic property of the seeds. variables were analysed through paired T test (Annexure 2) are summarized in Table 5. using SPSS. 3.4 Food formulation 3. RESULTS AND DISCUSSION Sensory evaluation was performed among 35 subjects with different categories of 3.1 Phytochemical analysis the food products. Statistical analysis was The Artocarpus hirsutus seeds are rich performed using paired sample‘t’ test among in phenolic compounds. The present study the different responses. The results revealed revealed the presence of ‘alkaloids and that the category C among all the foods tannins’ in aqueous extract, ‘alkaloids, tannins, (chapati, idly and steam cakes) was statistically sterols and quinine’ in methanol extract, and found to be more acceptable by the panel. ‘alkaloids and sterols’ in di-ethyl ether extract. Therefore, it can be concluded that higher The results obtained from the qualitative concentration of the seed powder phytochemical analysis are summarized in incorporated into the food products had better Table 1. sensory perception than the other concentrations.

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Table1. Qualitative (phytochemical) analysis of Artocarpus hirsutus seeds

Extracts Compounds Aqueous Methanol Di-ethyl ether Alkaloids + + + Terpenoids - - - Tannins + + - Sterols - + + Quinine - + - (+ present, - absent)

Table 2. Quantitative (nutrient) analysis of Artocarpus hirsutus seeds

Nutrient Optical Density value mg/g Carbohydrate 1.69 303 Protein 1.52 178 Sodium 0.039 28 Potassium 0.022 21 Calcium 0.032 26

Table 3. Percentage of Fibre, Moisture and Ash

Nutrient Percentage (%)

Fibre 13 Moisture 0.14 Ash 0.02

Table 4. Anti-bacterial and Anti-fungal activity – Zone of inhibition

Extracts Organism Aqueous Methanol Di-ethyl ether Disc DMSO

E.coli 9 mm 6 mm 3 mm 11 mm -

Staphylococcus aureus 11 mm 15 mm 3 mm 7 mm -

Klebsiella pneumoniae 10 mm 8 mm - 14 mm -

Salmonella typhi 14 mm 15 mm - 7 mm -

Aspergillus niger 5 mm 3 mm 5 mm -

Trichoderma viridae 2 mm 8 mm - -

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Table 5. Optical Density values for Anti-diabetic testing

Assay Optical Density Control Optical Density % of Value Value inhibition α-Amylase inhibition 0.029 0.631 95.4

α-Glucosidase 0.067 0.743 90.9 inhibition

Figure1. Zone of Inhibition of Artocarpus hirsutus seeds powder against bacterial and fungal strains

E. coli Staphylococcus aureus

Klebsiella pneumoniae Salmonella typhi

Aspergillus niger Trichoderma viridae

4. CONCLUSION Nutrient analysis shows that the seed powder In the current study, phytochemical is an excellent source of carbohydrate and analysis reveals the present of phenolic protein and it also has good anti-bacterial compounds which are known to exhibit action against food borne pathogens. The medicinal as well as physiological activities. study also reveals excellent anti-diabetic

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properties of the seeds. The study can be 8. Azeem AK, Rasheed A, Dilip C, Junise V, Rani S. further extended by conducting shelf-life and Diuretic activity of the of Artocarpus hirsutus Lam. J of Current Science 2011;1(1):16-9. cytotoxicity studies. Due to the anti-diabetic 9. Chowdhury AR, Bhattacharya A, Chattopadhyay P. properties, it can also be subjected to further Study on functional properties of raw and blended supplementation studies. Jackfruit seed flour (a non-conventional source) for food application. Indian J of Natural Products and REFERENCES Resources 2012;3(2):347-53. 1. Monish M, Sowmiya R, Ragunathan, Jesteena J. 10. Chun-Nan Lin, Chai-Ming Lu and Pao-Lin Huang. Extraction of Bioactive compounds from Cassia Flavonoids from Artocarpus heterophyllus. J Phyt auriculate pods and leaves and its medicinal uses. J of chem. 2005;39(6):1447-51. phytochemistry 2017; 6: 425-34. 11. Dibinital D, SathishSekar D, Senthil Kumar 2. Abedin MS, Nuruddin MM, AhmedKU, Hossain A. KL.Pharmacognostica Studies on the Bark of Nutritive compositions of locally available jackfruit Artocarpus hirsutus Lam. Hygeia J for Drugs and seeds (Artocarpusheterophyllus). International J of Medicines 2010;2(1):22-7. Biosciences 2002;40:1-7. 12. Downes FP, Ito K. Compendium of methods for the 3. Ahmedullah M, Nayar MP. Endemic of the microbiological examination of foods American th Indian region. American J of Clinical Nutrition Public Health Association. 4 ed. WashingtonDC; 1986;71(6):13-21. 2008:7-83. 4. AkhilHarietal .Artocarpus: A review of its phyto- 13. Elango R, BallRO,Pencharz PB. Amino acid chemistry and pharmacology. J of Pharma Search requirements in humans: with a special emphasis on 2014; 9(1)7-12. the metabolic availability of amino acids. J Biochem. 5. ArungET,Shimizu K, Kondo R. Artocarpus plants as a 2009;37(2):19-27. potential source of skin whitening agents Natural 14. Enos TA, Britanto DW, Yohana AH.Anti-cancerous Product Communications 2012;6(9):1397-402. properties of di-ethyl ether extract of from 6. Jayasri MA, Radha A, Mathew TL. Alpha-amylase and Sukun (Artocarpus altilis) in Human Breast Cancer alpha-glucosidase inhibitory activity of Costus pictus (T46D) cells Tropical. J of Pharmaceutical Research, Don in the management of Diabetes. J of Herbal 2009; 8(4):317-24. Medicine and Toxicology 2009;3(1):91-4 15. Fang et al., Anti-inflammatory effects of phenolic 7. Asha DS, Ben CP. Least concerned bark and stipules compounds isolated from the fruits of Artocarpus of Artocarpus species (Moraceae) - an effective anti- heterophyllus. J Agrc Food Chem 2008; 56(2):4463-8. bacterial agent. International Science Community Association 2014; 3(1):25-9.

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