Indian Journal of Traditional Knowledge Vol 18(1), January 2019, pp 16-24

Nutritional evaluation of Rhus chinensis Mill. (Heimang) and development of value added products

Memthoi Devi Heirangkhongjam*,1,2,+ & Iboyaima Singh Ngaseppam1,2 1Department of and Vegetable Technology, CSIR-Central Food Technological Research Institute, Mysuru 570 020, India. 2Academy of Scientific and Innovative Research (AcSIR), Ghaziabad 201 002, India. E-mail: [email protected]

Received 16 May 2018; revised 2 August 2018

Rhus chinensis Mill. is a lesser known highly acidic fruit, traditionally used for its digestive properties and treatment of diarrhea, dysentery and gastrointestinal ailments. In this study, physicochemical characteristics and nutritional properties of different parts of the fruit and development of value added products – R. chinensis tablet and R. chinensis candy and their storability were evaluated. Different parts of the fruit were found to have high content of total fibre-seed (25.97%), pulp (22.83%), whole fruit (22.15%), tablet (15.45%) and candy (14.47%), total fat-seed (36.96%), pulp (20.92%), whole fruit (12.97%), tablet (8.24%) and candy (6.13%) and total acidity-pulp (19.94%), whole fruit (7.28%), seed (3.12%), tablet (3.7%) and candy (3.68%). The major organic acids identified were malic acid, citric acid and ascorbic acid, which are documented for the first time. In comparison to whole fruit and seed, fruit pulp showed the maximum activities of antioxidants, total polyphenols and total flavanoids. These findings suggest that R. chinensis fruit can be considered as a potential functional food or functional food ingredient.

Keywords: Antioxidant, Functional ingredient, Medicinal , Organic acid, Traditional food IPC Code: Int. Cl.18 A61P 17/18, C04B 103/00, A61K 36/00, C08B 3/12

Fruits are not only rich in vital vitamins and minerals, full sun. The average height of the tree ranges from but being fibre rich can also support a healthy 12 to 15 ft and reaches upto 25 ft. The tree flowers digestive system. While in general are during summers and matures in October. It is also considered to be health protective foods, there are known as “September Beauty”. The tree bears several species that produce fruits with exceptionally odd-pinnately compound and creamy-white higher amounts of certain nutrients and, accordingly flowers. It bears tiny seeded fruits in a cluster with a may be more advantageous than others with regards waxy coating. The individual fruit weighing about specific dietary requirements. Despite their potential 16.23±2.66 mg is either orange or red in color at health benefits, such fruits often remain underutilized maturity and contains only one seed with a peculiar necessitating scientific evidence to convince and sharp acidic taste2,3. sensitize the people about their nutritional importance. Manipur has a rich diversity of wild indigenous Underutilized fruit crops are the type of plant species fruits identified in local communities through local conventionally used for food, fibre, fodder, oil or names by reason of their uses for food and medicine4,5 1 medicinal values . One such fruit, “Rhus chinensis and R. chinensis local name “Heimang” is one such Mill.” belonging to the family , fruit. It is an underutilized fruit due to its non common name “Nutgall tree” or “Chinese ” is a extensive use, but the fruit plays a major role in deciduous tree grown abundantly in , Japan and supplementing the diet of the local inhabitants. North eastern parts of India. It is widely found in However, the traditional herbal healers locally known Manipur state of North-East India, where it is as Maibas and Maibis and other traditional folklore of popularly called “Heimang”. The tree is favorably North-East India use the ripe fruits of R. chinensis for grown in warm temperate to tropical regions. It is best their supposed antiviral, antibacterial, anti-diarrhea, suited in well drained soil with acidic or alkaline in antioxidant and digestive properties6,7. It is also used ——————— in the treatment of intestinal worms, dyspepsia, and 8 *Corresponding author stomach ulcer, and as hair lotion . Nonetheless, HEIRANGKHONGJAM & NGASEPPAM: NUTRITIONAL EVALUATION OF RHUS CHINENSIS AND ITS 17 VALUE ADDED PRODUCTS review of literature indicates the lack of scientific fresh fruits are usually dried and stored to avoid data to support these nutritional and nutraceutical spoilage and future use. However, in order to properties. Moreover, use of R. chinensis fruits as accurately measure its nutritive value, the whole fruits commercial functional food or as food ingredients is were cleaned, dried at 55±5°C in a hot air oven. The virtually non-existent except limited use as an dried fruit was de-seeded in plate mill and passed acidulant in traditional fruit preparations. Therefore, through an aspirator for separation of dry skin cum this study was aimed to enumerate the nutritional pulp from the seeds. Both the fractions were stored in properties of R. chinensis fruit and to develop the airtight glass containers at 4°C until they were value-added products viz., R. chinensis tablet and required for the development of products as well as for R. chinensis candy with minimal expenses without analysis of physicochemical and nutritional properties. losing its taste and nutritional properties. R. chinensis tablet may be consumed as a mouth freshener and Chemicals digestive aid whereas R. chinensis candy as a high The following chemicals procured from Sigma- calorie and nutritious food. In fact, Allopathic Aldrich, Bangalore were used for different laboratory medicine can cure multiple diseases, but its determinations: Aluminium chloride, DPPH (2, 2- unregulated high prices and serious side effects are Diphenyl-1-picrylhydrazyl), gallic acid, Folin- letting many people to look for an alternative with Ciocalteu's reagent, Tripydyltriazine (TPTZ), ferric minimal cost and little or no side effects. Also about chloride (FeCl3 • 6H2O), rutin, malic acid, ascorbic 25 % of drug prescriptions are produced from natural acid, citric acid, fumaric acid and oxalic acid. 9 sources . Due to this, within the last couple of Preparation and evaluation of value-added products decades, the traditional medicinal /herbal The value added products of R. chinensis fruit were therapy has regained its prominence. Apart from formulated as per the procedure of Ayurvedic traditional medicinal use, some select high value Formulary of India14. medicinal plants act as an alternate source of income and employment generator for the local tribal R. chinensis tablet community in the region10. Moreover, Ministry of After trial experiments, fruit pulp powder (40%) was AYUSH and the government have undertaken mixed with other finely powdered ingredients namely several projects to start food industries and provide garlic (Allium sativum L., 17%), black pepper (Piper financial assistance to farmers to grow aromatic and nigrum L., 17%), black salt (9%), salt (4%) and sugar medicinal plants11,12. (13%) in fixed ratio to enhance the palatability and

Materials and methods

Authentication and plant material This study concentrates on the North-East India's traditional herbal healers' usage of R. chinensis. A field study was conducted to collect various information on traditional uses of R. chinensis according to the methodology of Jain13. For collection of the plant species, field visit was done accompanying local herbal practitioners at Bishnupur District, Manipur during October 2017. Then a voucher specimen of the plant was prepared and sent to Botanical Survey of India, Eastern Region Centre, Shillong, India. It was later verified and authenticated by Dr AA Mao, Scientist-F, Head of Department and Dr Chaya Deori, Scientist-D, Herbarium Incharge bearing accession number BSI/ERC/Tech/2017- 18/699 (Fig. 1).

Dried fruits are normally used as ingredients in local Fig. 1 — R. chinensis tree and different parts of R. chinensis fruit; culinary preparations instead of fresh fruits. Hence, the a. R. chinensis tree; b. whole fruit; c. pulp; d. seed. 18 INDIAN J TRADIT KNOWLE, JANUARY 2019

sensorial acceptability. In addition to this, 1% of guar method18. Total soluble solids (TSS, oBrix) content gum was added as a binding material, and tablets of was determined by using digital refractometer 10 mm diameter were made by using a hand operated (RX-5000 ATAGO) while pH was measured using mold. The prepared tablets were packed in an airtight pH tester_30 (Eutech Instruments, OAKTON). Total glass container and stored at room temperature titratable acidity (TTA%) of the samples was (25±5°C) to observe physicochemical changes. determined by titration method19, and expressed as anhydrous citric acid equivalent (% w/w). Reducing R. chinensis candy sugars were estimated by the Dinitrosalicylic Acid The candy was prepared by mixing the dried pulp method, and absorbance was read at 510 nm with powder (45%) with predetermined calculated 20 glucose as a standard for the calibration curve . quantities of other ingredients namely, jaggery (50%), chilli (0.4%), salt (4%), black salt (0.6%) and mixed Preparation of extracts with aqueous, methanol, thoroughly. In the preparation process, jaggery was ethanol, and acetone suspended in a little water in a thick round bottom Two g of the sample was extracted in 40 mL stainless stain vessel and boiled until melting into soft distilled water with 2 h intermittent shaking. The ball stage. The product mixture was then added to the solution was collected and centrifuged at 3,000 × g for boiling jaggery and continued heating with vigorous 10 min. Then, the filtrate was concentrated in rotary stirring till the product become a soft lump. The vacuum evaporator at 45°C for 20 min at 70 rpm, and prepared lump was then poured into a greased the concentrated residue was dissolved in 20 mL stainless steel plate, molded into the desired shapes distilled water and finally stored at 4°C. The same and sizes and stored in an airtight glass container and procedure was carried out for other solvents (80% stored at room temperature to observe methanol, ethanol, and acetone). physicochemical changes. Determination of total polyphenols Microbial analysis of value added products The Folin–Ciocalteu reagent assay was used for The product samples were serially diluted (10-1 to determining the total phenolics content according to 10-5) in 9 mL of saline. Surface plating with the method of Singleton & Rossi21. An aliquot of the appropriate dilution was carried on to Violet Red Bile samples was mixed with 5 mL Folin–Ciocalteu Agar and Rose Bengal Chloramphenicol for coliform reagent and 4 mL sodium carbonate (7.5%). The and yeast and mold, respectively and incubated absorbance was read at 765 nm, and gallic acid was anaerobically at 37 °C for 48 h. Total viable counts used as a standard for the calibration curve. The total were determined using plate count agar incubated at amount of phenolics compounds was calculated and 25°C for 24 h15. expressed as gallic acid equivalent (mg/100 g).

Sensory evaluation of value-added products Determination of total flavanoids content To determine the consumer preferences and The determination of flavanoids was performed acceptability of the R. chinensis based products, sensory using modifications according to the colorimetric assay evaluation was carried out using a five Point Hedonic 22 16 of Kim et al. . An aliquot of the samples was mixed Scale , with 1-dislike very much, 2-dislike moderately, with 5% sodium nitrite solution, incubated at ambient 3-neither like nor dislike, 4-like moderately and temperature for 5 min, and then 10% aluminium 5-like very much. Thirty semi-trained panelists, aged chloride solution was added, followed by 2 mL of 1M (25-35 yrs) familiar with the sample, working at sodium hydroxide. The absorbance was monitored at Department of Fruit and Vegetable Technology, CSIR- 17 510 nm. A calibration curve was prepared with Rutin, CFTRI, Mysuru carried out the sensory evaluation . and the results were expressed as Rutin (mg/100 g). The sensory attributes assessed were appearance, aroma, taste, mouth feel, and overall acceptability. Determination of antioxidant properties by ferric reducing/antioxidant power (FRAP) method Physicochemical analysis and nutritional properties The antioxidant capacity of the extract was Proximate composition and chemical analysis determined using a modification of the ferric The proximate composition was estimated reducing/antioxidant power (FRAP) assay of Benzie & according to the method of Rangana17. The per cent Strain23. Three mL of FRAP reagent, prepared freshly carbohydrates content was calculated by difference and warmed at 37°C, was mixed with 40 μL of plant HEIRANGKHONGJAM & NGASEPPAM: NUTRITIONAL EVALUATION OF RHUS CHINENSIS AND ITS 19 VALUE ADDED PRODUCTS extract and reaction mixtures were incubated at 37°C using 5-Point Hedonic Scale for the sensory attributes and absorbance was read at 593 nm. Aqueous solutions like appearance, aroma, taste, mouth feel and overall of known Fe (II) concentrations in the range of 100– acceptability. The results are presented in Fig. 3. Both 2000 μM (FeSO47H2O) were used for calibration. the products received overall acceptability by the panelists. Determination of antioxidant properties by DPPH radical scavenging assay method Proximate composition and chemical analysis The antioxidant property of the extract was Proximate compositions of different fruit portions estimated according to the method of Reddy et al.24. were determined. The results are summarized in An aliquot of the sample (0.1-0.5 mL) was mixed Table 1. Crude fibre and fat contents in different with 2.9 mL DPPH solution (0.1 mM in methanol) fractions/products decreased in the order, seed > pulp and allowed to stand for 20 min at RT (dark). After > whole fruit > tablet > candy. TTA (%) was found to which the absorbance was read at 517 nm against the be highest in the pulp (19.94%) than whole fruit, seed methanol as blank and methanol + 2.9 mL DPPH as and value added products. the control. Analysis of total polyphenols, total flavanoids and Organic acid determination by HPLC method antioxidant activities

Preparation of extracts The polyphenols content was found to be Fruit segments and value-added samples were significantly more in the pulp as compared to the suspended in water in the ratio of 1:10 (w/v), whole fruit and seed. It was also observed that 80% centrifuged at 3000 x g for 15 min. The filtrate was ethanol extract contained highest polyphenols in pulp concentrated in rotary vacuum evaporator at 45°C for (6283.33 GAE mg/100 g). Whereas, acetone extracts 20 min at 70 rpm, and the dried residue was dissolved showed highest polyphenols in whole fruit (4256.66 in 10 mL distilled water and filtered through a 0.45 GAE mg/100 g) and seed (327.42 GAE mg/100 g). In µm membrane filter and injected directly into HPLC. candy and tablet, methanol extract showed the highest

Chromatographic conditions Stock solutions of oxalic, citric, malic, ascorbic and fumaric acids were prepared in millipore water and made into appropriate dilutions. Shimadzu Class-10 VP HPLC system with diode-array detector apparatus was used with a UV absorbance detector set at 215 nm. The chromatographic separation was performed on Ascentis C-18, 5 µm (25cm × 4.6mm). The organic acid was determined using a modification of Tomlins et al.25. Identification of chromatographic peaks was Fig. 2 — Value added products of R. chinensis fruit; a. candy; b. tablet. done by comparing the retention times of the samples with those of the standards. The quantities of organic acids were estimated from the peak areas by injecting the known amounts of the standards.

Statistical analysis All determinations were conducted in triplicate and subjected to single ANOVA and multiple comparison test was done by Tukey’s test using Graph Pad Prism software.

Results

Sensory evaluation of value-added products The prepared value added products from R. chinensis pulp (Fig. 2) were subjected to sensory evaluation Fig. 3 — Sensory quality scores of R. chinensis value added products 20 INDIAN J TRADIT KNOWLE, JANUARY 2019

Table 1 — Proximate composition and chemical analysis of different parts of R. chinensis fruit and its value added products Parameters Whole fruit Pulp Seed Candy Tablet Moisture (%) 7.1±0.2a 4.26±0.2a 4.5±0.30a 1.36±0.06b 5.21±0.59a Crude Fat (%) 12.97±0.87c 20.92±4.01b 36.96±1.03a 6.13±1.3d 8.24±0.38d Ash (%) 2.30±0.1c 3.83±0.1c 3.22±0.08c 17.88±0.25b 18±0.91a Crude fiber (%) 22.15±0.1a 22.83±0.6a 25.97±2.05a 14.47±0.55c 15.45±0.29b Crude Protein (%) 6.12±0.4b 4.25±0.2b 13.09±0.61a 4.59±0.11b 4.47±0.31b Carbohydrate (%) 49.35±1.16b 43.89±3.04b 16.24±1.05c 55.57±1.13a 49.44±1.52b Reducing sugar (%) 1.27±0.47b 1.92±0.70b 1.02±0.36b 8.10±0.67a 3.30±0.56b Total Soluble Solids (oBrix) 13.66±1.52d 30.1±0.3c 20.33±1.52d 79.33±0.30a 62.6±0.2b Acidity (%) as anhydrous citric acid 7.28±0.47b 19.94±0.47a 3.12±0.62c 3.68±0.04c 3.7±0.08c pH 3.44a 3.19a 4.21a 3.32a 3.5a * Values are given in mean ± standard deviation of three independent determinations. Values in the same row followed by the same letter are not significantly different at p ≤ 0.05 as measured by Tukey’s test.

Table 2 — Total polyphenol content (TPC) and Total flavanoids content (TFC) of different parts of R. chinensis fruit and its value added products Sample TPC (GAE mg/100g extract) TFC (Rutin mg/100g extract) Aqueous Acetone Ethanol Methanol Aqueous Acetone Ethanol Methanol Whole fruit 725± 4256.66± 2836.66± 3576.33± 57.33± 644.18± 310.6± 524.64± 2.64db 23.71aa 45.09cb 171.20bb 1.13db 4.78ab 2.69cb 17.68ba Pulp 2028.33± 4506.66± 6283.33± 6143± 85.39± 709.77± 495.29± 554.32± 18.58da 179.25ca 267.31aa 20.66ba 4.57da 20.12aa 1.98ca 15.72ba Seed 271.25± 2829.58± 1163.95± 829.78± 52.46± 327.42± 309.18± 174.87± 9.94dd 127.67ab 39bd 33.09cd 1.79cb 16.40ac 19.53ab 19.40bb Candy 810.57± 1086.75± 1116.64± 1560.83± 35.67± 196.11± 129.0± 202.89± (zero day) 34.97cb 126.73bd 41.36bd 99.23ac 3.90cc 8.65ad 7.78be 17.40ab Candy 796.40± 1048.01± 1092.86± 1520.86± 34.41± 186.57± 124.25± 189.62± (30 days) 26.24cb 147.27bd 46.91bd 113.62ac 3.17cc 9.08ad 8.63be 9.09ab Candy 760.01± 952.56± 1070.21± 1486.39± 25.65± 165.90± 118.26± 170.86± (60 days) 33.27cb 12.25bd 43.92bd 117.87ac 3.40cd 12.76ad 7.56be 9.68ab Candy 751.82± 926.16± 1048.99± 1437.46± 22.27± 146.07± 109.05± 156.09± (90 days) 18.50cb 14.07bd 45.57bd 130.45ac 2.61cd 18.87ad 6.29be 20.30ac Tablet 485.93± 1461.94± 1355.60± 1571.06± 45.47± 349.96± 209.57± 152.06± (zero day) 12.52bc 72.03ac 51.87ac 108.17ac 2.57dc 35.15ac 8.85bc 3.46cc Tablet 460.83± 1425.45± 1308.13± 1536.11± 41.71± 319.93± 193.07± 141.34± (30 days) 5.51bc 85.42ac 82.26ac 86.93ac 3.00dc 15.31ac 0.64bc 3.41cc Tablet 429.65± 1403.77± 1258.62± 1491.98± 39.07± 278.64± 162.03± 122.36± (60 days) 5.61bc 67.58ac 20.14ac 60.70ac 2.96dc 6.99ac 19.85bd 8.46cd Tablet 408.33± 1347.78± 1221.49± 1449.90± 35.38± 259.0± 140.60± 104.10± (90 days) 15.11bc 118.31 ac 41.65ac 54.78ac 2.71dc 4.36ac 14.30bd 8.72cd * Values are given in mean ± standard deviation of three independent determinations. Values in the same row (first letter- comparison of same sample in different solvent extracts) and column (second letter- comparison of different samples in same solvent extract) followed by the same letter are not significantly different at p ≤ 0.05 as measured by Tukey’s test. activities of 1560.83 and 1571.06 GAE mg/100 g, 709.77, 644.18, 349.96, 327.42 and 196.11 GAE respectively in zero day study. However, over 90 days mg/100 g, in pulp, whole fruit, tablet (zero day), seed study period there was no significant changes in the and candy (zero day), respectively, as compared to the extracts (Table 2). Similarly, total flavanoids content other extracts. Also, no significant changes were was found significantly higher in pulp compared to observed over 90 days storage period (Table 2). whole fruit, seed and value added products. The Furthermore, decrease in total polyphenols and acetone extract had the highest flavanoids content of flavonoids content were observed during processing HEIRANGKHONGJAM & NGASEPPAM: NUTRITIONAL EVALUATION OF RHUS CHINENSIS AND ITS 21 VALUE ADDED PRODUCTS in comparison to the pulp. In candy, 59% and 38% extract of candy at zero day (3260.07 mM/100 g) and change was observed in polyphenols and flavanoids acetone extract of tablet at zero day (3248.55 mM/100 g) whereas, 35% and 23% change was observed in (Table 3). Moreover, decrease in scavenging activity tablet respectively. was observed during the storage period with 11 % and DPPH assay is one of the most widely used 7% in tablet and candy respectively. methods for screening antioxidant activity of plant extracts26. Radical scavenging percentage was Organic acid determination of different fruit parts measured to assay the antiradical activity of the In the present study, we observed a high titratable fruit extracts. The antioxidant activity was found acidity in R. chinensis fruits. Therefore, extracts of R. to be more in acetone extract of the pulp (9.31 chinensis whole fruit, pulp, seed and value-added IC50 µg/mL) followed by whole fruit, seed, tablet products were subjected to HPLC analysis for (zero day) and candy (zero day) with 10.35, 14.65, determining organic acid content which is documented 46.67 and 109.92 IC50 (µg/mL), respectively. Also, for the first time. The content of malic acid in pulp no significant changes were observed over the (12.27 g/100 g) was found to be significantly high storage period (Table 3). Different extracts being compared to whole fruit (7.16 g/100 g), seed (3.43 rich in total phenolics have the potential of g/100 g), candy at zero day (8.74 g/100 g) and tablet at scavenging free radicals and hence protecting human zero day (6.79 g/100 g). Citric acid content in pulp, body from certain ailments. whole fruit, seed, candy (zero day) and tablet (zero In FRAP, the highest scavenging activity was found day) was 7.67 g/100 g, 4.62 g/100 g, 3.87 g/100 g, in the ethanolic extract of pulp (14521.41 mM/100 g), 5.07 g/100 g and 5.33 g/100 g, respectively. The compared to acetone extracts of whole fruit (7292.31 values of ascorbic, oxalic and fumaric acids also mM/100 g) and seed (3726.83 mM/100 g), methanol varied in different fruit parts and processed products

Table 3 — DPPH and FRAP of different parts of R. chinensis fruit and its value added products

Sample DPPH IC50 (µg/mL) FRAP (mM/100 g extract) Aqueous Acetone Ethanol Methanol Aqueous Acetone Ethanol Methanol Whole fruit 86.54± 10.35± 11.19± 12.27± 1715.74± 7292.31± 7011.88± 6393.38± 0.64ba 0.13aa 0.22aa 0.04aa 14.50dc 99.36ab 152.45bb 89.51cb Pulp 84.47± 9.31± 9.55± 11.64± 3740.27± 9216.96± 14521.4± 9366.79± 0.59ba 0.06aa 0.10aa 0.16aa 32.76ca 66.53ba 96.66aa 46.57ba Seed 116.37± 14.65± 18.87± 23.35± 522.69± 3726.83± 2630.05± 2038.44± 2.58bb 0.15aa 0.2ab 0.10ab 27.78de 43.63ac 67.93bc 27.23ce Candy 267.15± 109.92± 98.78± 78.25± 2176.74± 3046.86± 2151.77± 3260.07± (zero day) 0.43bd 0.20ac 0.38ad 0.10ac 28.42cb 38.36bf 20.77ce 20.23ac Candy 272.67± 118.94± 110.69± 83.75± 2092.22± 3029.58± 2145.17± 3182.24± (30 days) 3.10dd 0.46cc 0.38bd 0.36ac 53.54db 54.46bf 52.81ce 25.11ac Candy 283.76± 118.96± 106.53± 89.88± 2036.52± 2927.77± 2067.25± 3127.54± (60 days) 11.05dd 0.13cc 1.65bd 0.65ac 9.98cb 20.23bg 31.73cf 14.50ac Candy 294.69± 139.29± 118.97± 94.10± 1957.77± 2864.39± 1971.21± 3099.88± (90 days) 2.01dd 1.40cd 1.07bd 0.86ac 14.50cb 8.80bg 11.99cf 34.73ad Tablet 194.20± 46.67± 67.06± 61.03± 1170.23± 3248.55± 2578.19± 2157.53± (zero day) 0.24cc 0.28ab 0.40bc 0.1bc 17.60dd 29.0ad 35.05bd 23.04ce Tablet 199.49± 69.48± 85.15± 65.69± 1120.29± 3192.84± 2549.37± 2117.19± (30 days) 1.10cc 0.43ab 0.64bd 0.29ac 15.24dd 44.01ae 49.23bd 11.52ce Tablet 209.93± 70.58± 96.18± 75.94± 1072.27± 3069.91± 2432.20± 2030.76± (60 days) 0.65cc 0.47ab 4.02bd 0.49ac 8.80dd 14.50af 14.50bd 15.24ce Tablet 212.77± 74.47± 106.98± 85.97± 970.47± 2952.74± 2341.93± 1969.29± (90 days) 0.59dc 1.21ab 1.28cd 0.1bc 23.04dd 11.52ag 15.24bd 14.50ce * Values are given in mean ± standard deviation of three independent determinations. Values in the same row (first letter- comparison of same sample in different solvent extracts) and column (second letter- comparison of different samples in same solvent extract) followed by the same letter are not significantly different at p ≤ 0.05 as measured by Tukey’s test. 22 INDIAN J TRADIT KNOWLE, JANUARY 2019

Table 4 — Organic acid determination of different parts of R. chinensis fruit and its value added products by HPLC method Sample Oxalic acid (g/100 g) Malic acid (g/100 g) Ascorbic acid (g/100 g) Citric acid (g/100 g) Fumaric acid (g/100 g) Whole fruit 0.13±0.004ca 7.16±1.54ab 0.41±0.12ca 4.62±0.46bb 0.02±0.003ca Pulp 0.15±0.004ca 12.27±0.17aa 0.57±0.001ca 7.67±0.53ba 0.03±0.01ca Seed 0.14±0.001ba 3.43±0.09ac 0.29±0.01ba 3.87±0.36ab 0.02±0.001ba Candy (zero day) 0.40±0.003ca 8.74±0.14ab 0.17±0.004ca 5.07±0.14bb 0.04±0.00ca Candy (30 days) 0.30±0.01ca 7.52±0.25ab 0.17±0.001ca 4.59±0.21bb 0.03±0.001ca Candy (60 days) 0.22±0.001ba 5.61±0.61ab 0.15±0.003ba 4.46±0.02ab 0.02±0.001ba Candy (90 days) 0.24±0.03ba 3.25±0.72ac 0.14±0.002ba 3.60±0.002ab 0.02±0.01ba Tablet (zero day) 0.25±0.01ca 6.79±0.47ab 0.37±0.002ca 5.33±0.21bb 0.03±0.001ca Tablet (30 days) 0.25±0.001ca 6.80±0.42ab 0.32±0.02ca 4.04±0.22bb 0.03±0.001ca Tablet (60 days) 0.24±0.01ca 6.21±0.09ab 0.28±0.003ca 3.72±0.11bb 0.02±0.001ca Tablet (90 days) 0.16±0.001ba 4.11±0.36ac 0.22±0.02ba 3.25±0.28ab 0.02±0.001ba * Values are given in mean ± standard deviation of three independent determinations. Values in the row (first letter- comparison of different organic acid in same sample) and column (second letter- comparison of same organic acid in different sample) followed by the same letter are not significantly different at p ≤ 0.05 as measured by Tukey’s test.

(0.57-0.02 g/100 g). While both malic acid and citric TSS (%), total fat content, etc also increases. Hence, acid decreased in candy and tablet, no changes were due to these changes the present findings and previous observed in ascorbic, fumaric and oxalic acid contents reports were found to be different or similar. Since the (Table 4). fruit pulp contains high acid, it can be used as an acidulant (functional ingredient) in the preparation of Discussion many food products. R. chinensis fruit is a highly acidic fruit and hence Dried pulverized whole fruit, pulp and seed of usually not consumed fresh. While preparing the R. chinensis in four different solvents-aqueous, value added products, necessary care was taken to acetone, ethanol and methanol were used as extracts. select right type of ingredients (e.g., garlic, black The (abnormal outgrowth of the plant caused by pepper, black salt, sugar, guar gum, jaggery, chilli, infestation) of R. chinensis, considered to and salt) in a proper ratio to ensure that final products possess many natural antioxidants, have attracted were palatable and retained the desired sensorial interest because of their protective and potential properties. The choice of fruit pulp for value addition therapeutic effects29. Many phenolic compounds was based on its higher fibre content, titratable have antioxidant properties, which may contribute acidity, organic acid and antioxidant properties than to protection against oxidative damage30. According whole fruit and seed. Microbiological analysis to the reports of Cai et al.31, the total polyphenol confirmed that R. chinensis products (candy and content of aqueous and methanol extract of tablet) were free from yeast, mold and pathogenic R. chinensis fruit was found to be highest of all microorganism (Escherichia coli) and thus safe from 112 Chinese medicinal herbs. Similar, antioxidant the consumer health standpoint. The nutritional activity was reported by Sharma et al.27. A higher properties of different parts of the fruit and value DPPH value of the extracts, implies their strong added products were also determined with a view to antioxidant potential. provide scientific evidence that could increase their The free radical scavenging activity was acceptability and consumption. The crude fiber determined by FRAP method, there was an increase in content of the pulp was found within the range the absorbance of extracts in progressive manner reported by Sharma et al.27. However, percentage of because of elevated ferric reducing power with total fat observed in this study differed with the increased concentrations. Therefore, all the extracts reports of Sharma et al.27 but was in agreement with (pulp, whole fruit, seed, candy and tablet) can donate the findings of Chen et al.28. Similarly, TTA (%) of electrons to free radicals which stably exists our fruit (16.51%) was also similar to the findings of biological system or generated through consumption Sharma et al.27. With the extension in storage period, of various food products, and hence confirmed that the physicochemical characteristics, viz. TTA (%), R. chinensis fruit is an antioxidant rich source. HEIRANGKHONGJAM & NGASEPPAM: NUTRITIONAL EVALUATION OF RHUS CHINENSIS AND ITS 23 VALUE ADDED PRODUCTS

Traditional significance of the study to the Shirui- Khangkhui and Kailam. Also, the government society/researchers had established two Herbal Gardens for ex-situ Since Manipur is located in one of world's bio- conservation of medicinal plants at Langol and Raj diversity hotspot "Indo-Burma" region, it has rich bio- Bhawan with financial assistance from the National diversity. It is reported that around 1200 medicinal Medicinal Plants Board, Ministry of Health & Family plants, indicating 430 species with local traditional Welfare (now Ministry of AYUSH), Government of medicinal uses are found in the state. R. chinensis is India. Moreover, traditional knowledge of people in one such traditional medicinal species found in the local communities and establishing sacred grooves are region abundantly and with its long history of some of the ways of conserving genetic diversity of medicinal usage, the plant could potentially be a game medicinal plants in the natural habitats and changer for the local tribal community if these value- environment in relation to endangered species. added products achieve commercial success. As these value added products are 100% natural and highly Conclusion effective, it has good scope of replacing the currently R. chinensis fruit was found to have high amounts commercially available digestive pharmaceutical of crude fibre, crude fat and high total titratable products which has major side effects on human acidity. The major organic acids detected were malic, health. With unimaginable side effects of citric and ascorbic acid. Furthermore, the pulp was pharmaceutical products, there has been a tremendous found to have higher antioxidant properties compared rise in dependency on herbal and naturally produce to the whole fruit and seed. Decreases in total acids, medicines all over the world. As per the report, phenolics and antioxidant values were observed in around 80% of the population relies on traditional candy and tablet. Also, methanol and acetone extracts medicine. Hence, this dependency could be harnessed were found to be better compared to various solvents in generating employment and extra income for the extraction. The high phenolics and antioxidant local women and youth with the establishment of contents may have a significant role in the treatment small cottage based industries and thereby improving of diseases. Therefore, it could potentially be their livelihood. considered as a functional food or functional food For promoting the medicinal plants sector, the ingredient. Although R. chinensis and similar plant Government of India has set up National Medicinal species are well known for their traditional values, Plants Board (NMPB) on 24th November 2000. Its detailed scientific investigations are necessary to primary objective is to develop an appropriate ensure their sustainable commercial applications. mechanism for coordination among different ministries Conflict of interest for conservation, cultivation, trade and export by The authors have no conflict of interest to disclose. formulating policies and programme for the growth of medicinal plants sector at the State, Central and References International level. Moreover, the Government of India 1 Adiyaman P, Kanchana S, Usharani T, Ilaiyaraja N, had recently introduced "Ministry of AYUSH" as the Kalaiselvan A & Kumar AK, Identification and separate nodal ministry highlighting the growing quantification of polyphenolic compounds in underutilized fruits (Star fruit and Egg fruit) using HPLC, Indian J Tradit importance of herbal medicine in India and the world. Knowle, 15 (3) (2016) 487-493. Manipur being the gateway to South East Asia, 2 Miller AJ, Young DA & Wen J, Phylogeny and there is tremendous potential in promoting biogeography of Rhus (Anacardiaceae) based on ITS entrepreneurship in Aromatic & Medicinal plants. sequence data, Int J Biol Sci, 162 (6) (2001) 1401-1407. Therefore, in order face the challenges of over- 3 Tianlu M & Barfod A, Rhus Linnaeus, Flora China, 11 (2008) 335. exploitation of these medicinal plants because of 4 Samuel BO, Peter TM & Peter TJ, Detecting threats to commercial purposes, M/S Manipur Food Industries Encephalartos transvenosus (Limpopo cycad) in Limpopo Corporation, an undertaking of the Department has province, South Africa through indigenous knowledge, been working out a model where farmers grow Indian J Tradit Knowle, 16 (2) (2017) 251-255. aromatic and medicinal plants in their own land with 5 Sinha SC, Medicinal Plants of Manipur, (Manipur Cultural Integration Conference, Imphal, India), 1996. financial assistance from banks and supplying 6 Singh HB, Sandhu RS & Singh JS, Herbal Medicine of planting materials. Currently, the state has three Manipur: A Colour Encyclopaedia, (Daya Publishing House, Medicinal Plants Conservation Areas – Langol, New Delhi), 2003. 24 INDIAN J TRADIT KNOWLE, JANUARY 2019

7 Singh SR, Phurailatpam AK, Wangchu L, Ngangbam P 21 Singleton VL & Rossi JA, Colorimetry of total phenolics & Chanu TM, Traditional medicinal knowledge of with phosphomolybdic-phosphotungstic acid reagents, underutilized minor fruits as medicine in Manipur, Am J Enol Vitic, 16 (3) (1965) 144-158. Int J Agric Sci, 4 (8) (2014) 241-247. 22 Kim DO, Jeong SW & Lee CY, Antioxidant capacity of 8 Khumbongmayum AD, Khan ML & Tripathi RS, phenolic phytochemicals from various cultivars of plums, Ethnomedicinal plants in the sacred groves of Manipur, Food Chem, 81 (3) (2003) 321-326. Indian J Tradit Knowle, 4 (1) (2005) 21-32. 23 Benzie IF & Strain JJ, Ferric reducing/antioxidant power 9 Turker AU, Arzu BY, Fatma PK & Hakan T, In vitro assay: Direct measure of total antioxidant activity of antibacterial and antitumor efficiency of some traditional biological fluids and modified version for simultaneous plants from Turkey, Indian J Tradit Knowle, 17 (1) (2018) measurement of total antioxidant power and ascorbic acid 50-58. concentration, Methods Enzymol, 299 (1999) 15-27. 10 Kala CP, Medicinal plants conservation and enterprise 24 Reddy CVK, Sreeramulu D & Raghunath M, Antioxidant development, Med Plants, 1 (2) (2009) 79-95. activity of fresh and dry fruits commonly consumed in India, 11 Anonymous, National Medicinal Plants Board, (Ministry of Food Res Int, 43 (1) (2010) 285-288. AYUSH, Government of India), 2016. 25 Tomlins K I, Baker DM & McDowell IJ, HPLC method for 12 Imphal Free Press, Directorate of Information & Public Relation the analysis of organic acids, sugars, and alcohol in extracts (DIPR), (City Convention Centre, Imphal, India), 2015. of fermenting cocoa beans, Chromatographia, 29 (11) (1990) 13 Jain SK, A Manual of Ethno-botany, (Scientific Publishers, 557-561. Jodhpur, India), 1987. 26 Nanjo F, Goto K, Seto R, Suzuki M, Sakai M & Hara Y, 14 Anonymous, The Ayurvedic Formulary of India, Part I, 2nd Scavenging effects of tea catechins and their derivatives edn, (Government of India, Ministry of Health and Family on 1, 1-diphenyl-2-picrylhydrazyl radical, Free Radic Bio Welfare, New Delhi), 2003. Med, 21 (6) (1996) 895-902. 15 Moyane JN & Jideani AI, The physicochemical and 27 Sharma PB, Handique PJ & Devi HS, Antioxidant properties, sensory evaluation of commercial sour milk (amasi) physico-chemical characteristics and proximate composition products, Afri J Food Sci, 7 (4) (2013) 56-62. of five wild fruits of Manipur, India, J Food Sci Technol, 52 16 Tolia V, Han C, North JD & Amer F, Taste comparisons (2) (2015) 894-902. for lansoprazole strawberry-flavoured delayed-release orally 28 Chen CW, Zhang L, HE XM, Yang L & Jing, SB, disintegrating tablet and ranitidine peppermint-flavoured Convention Nutritional Components Analysis on Fruits of syrup in children, Clin Drug Investig, 25 (5) (2005) 285-292. R. chinensis Mill., Anim Husbandry Feed Sci, 4 (2010) 004. 17 Ranganna S, Handbook of analysis and quality control for 29 Rufino MS, Alves RE, Fernandes FA & Brito ES, Free fruit and vegetable products, (New Delhi: Tata McGraw-Hill radical scavenging behavior of ten exotic tropical fruits Publishing Company Ltd), 1986. extracts, Food Res Int, 44 (7) (2011) 2072-2075. 18 Gopalan C, Sastri BVR, & Balasubramanian SC, Nutritive 30 Blokhina O, Virolainen E & Fagerstedt KV, Antioxidants, value of Indian foods, (National Institute of Nutrition, oxidative damage and oxygen deprivation stress: a review, ICMR, India), 2007. Ann Bot, 91 (2) (2003) 179-194. 19 AOAC, Official Methods of Analysis, (Association of Official 31 Cai Y, Luo Q, Sun M & Corke H, Antioxidant activity and Analytical Chemists, Gaithersburg, MD, USA), 2000. phenolic compounds of 112 traditional Chinese medicinal 20 Sadasivam S & Manickam A, Biochemical methods, plants associated with anticancer, Life Sci, 74 (17) (2004) (New Age International (P) Ltd, Delhi), 1991. 2157-2184.