Appendix IV IERP PROJECT ANNUAL PROGRESS REPORT 2016-17 Period from 1st April 2016 to 31st March 2018 1. Project Title: “Ecological, Taxonomical and Ethnobotanical Study of Wild Edible Plants of Nainital District in Uttarakhand, India” 2. Name of Principal Investigator Dr. Lalit M. Tewari, Professor, Department of Botany, D. S. B. Campus, Kumaun University Nainital 263002; Uttarakhand, India Email: [email protected] Co- Principal Investigator(s) i) Dr. Ashish Tewari Assistant Professor Department of Forestry & Environmental Sciences D. S. B. Campus, Kumaun University, Nainital , 263002, Uttarakhand, India Email: [email protected] ii) Dr. Geeta Tewari Assistant Professor Department of Chemistry D. S. B. Campus, Kumaun University, Nainital, 263002, Uttarakhand, India Email: [email protected] Project staff Neha Chopra, JPF, Research Scholar, Department of Botany, D. S. B. Campus, KU Nainital 3. GBPIHED project sanction letter no. & Date of sanction: Letter No. GBPI/IERP- NMHS/16-16/48/14 dated: 31st March 2016 4. Total outlay sanctioned- Rs. 13, 60000/-(Thirteen lakh sixty thousand only) 4.1 Duration: 3 years 5. Date of start: 1st April 2016 6. Grant received during the year: 2016-17 Rs. 4, 97,000/- (Four lakh ninety seven thousands) 1 7. Expenditure incurred during the year: 2017-18 Copy of utilization certificate is attached with progress report as Annexure I. 8. Broad area of research: Ecological, Taxonomical and Ethnobotanical Study of Wild Edible Plants 9. Sub area of the project: Distribution, and nutrient analysis of wild edible plants 10. Approved objectives of the project: 1. Identification and taxonomic description of all wild edible plants distributed in the study areas with their relevant names and habitat distribution. 2. Develop an in-depth understanding of the role of ethnobotanical and indigenous knowledge about wild plant identification, collection, and usage in improving health with attention to the recording and dissemination of traditional methods of identification, sustainable harvesting, and consumption. 3. To assess the habitat, ecology, diversity, regeneration status, population and fruiting pattern of some selected edible wild plants of Nainital district of Kumaun. 4. Evaluate the nutritional value and variants of most preferred and important wild edible plants. 5. Prioritize important species of wild edibles and popularize them with the community. 6. Preservation of plant specimens and herbarium development will be done which will help in their easy identification in the field. 7. Undertake a program of promotion of wild plants to improve dietary diversity at local levels and evaluate the impact of the program on the improvement of economic and health conditions of the rural poor 11. Methodology 11.1 Study Area Present study was done for sub-tropical forest of Nainital district (Fig. 1 and Photoplate 1). Wild edible plants were collected from Ramnagar, Betalghat, Bhowali, Birla Chungi, Shyamkhet, China peak, Mangoli, Jeolikot and Ramgarh (Photoplate 1). 2 Fig 1: Map of Nainital District (http://www.uttaranchal.org.uk) 3 Ramnagar Betalghat Mangoli Bhowali Shyamkhet Jeolikot China Peak Birla Chungi Ramgarh Photoplate 1: Photographs of sites 4 11.2 Collection and Identification of Plants The study is based on ecology, taxonomy and ethnobotanical study of wild edible plants. Regular field surveys were conducted during 2016-2018 to document the wild edible plants. Specimens of all the plants were collected, preserved and identified by Prof. Y. P. S. Pangtey, Department of Botany, D. S. B. Campus, Kumaun University, Nainital. The well preserved plant specimens were deposited in the Department of Botany, D. S. B. Campus Kumaun University Nainital. Photographs of some important specimen have been taken for further details (Photoplate 2). The collection of wild edible plants was recorded with the help of local/ rural peoples and the information regarding local names, edible part, mode of use and ethnobotanical importance of wild edible was gathered. The information was collected by the observations and semi structured interview method with the people of different genders and age groups. 11.3 Assessment of habitat ecology, population study, plant diversity and regeneration status The field surveys were conducted during the year 2016-2018 within the selected sites for the assessment of vegetation. The vegetational data was analyzed for frequency, density, abundance (Curtis and McIntosch, 1950), relative frequency, relative density and relative basal area. In each site, Trees were sampled by randomly laid 30, 10x10m quadrat; shrubs by 30, 5x5m quadrat and herbs by 30, 1x1m quadrat in each site. For the collection of data from these quadrats standard ecological methods (Saxena & Singh, 1982; Dhar et al., 1997; Joshi & Samant, 2004; and Samant & Joshi, 2005) were followed. The circumference at breast height (cbh at 1.37m from ground) for each tree individual was recorded. Communities were identified based on the Importance Value Index (IVI). The IVI was calculated as sum of relative frequency, relative density and relative basal area. The abundance data of different sites were pooled to get community averages in terms of Density, Total Basal Area and IVI. In order to develop population structure and to understand regeneration of different species, individuals were measured for circumference at breast height (cbh) with a girthing tape, at each site all individuals were counted for each tree species. In addition to seedling and sapling classes (Good and Good, 1972), six or more classes based on cbh were arbitrarily established as follows: A. Seedlings < 10 cm cbh; B. Saplings 10~30 cm cbh; C. Young trees 31~60 cm cbh; 5 D. Pole size trees 61~90 cm cbh; E. Mature trees category I 91~120 cm cbh; F Mature trees category II 121~150 cm cbh; G Old trees >151cm cbh; and 11.4 Evaluation of the nutritional value and antioxidant activity of most preferred and important wild edible plants Some of the preferred and important wild edible plants (Table A) were brought to the laboratory and oven dried at 45o C. The dried edible parts were crushed and grind to obtain fine powdered form, which was further used for nutrient analysis (Protien, ash, total phenolic content, tannins, carbohydrate, sugar, metals). Table A: Plant part used for nutrient analysis Botanical Name Part used for analysis Carissa congesta Fruit Cinnamomum tamala Leaf Diplazium esculentum Aerial part Emblica officinalis Fruit Murraya koengii Leaf Rumex hastatus Aerial part Terminalia chebula Fruit Viburnum mullaha Fruit Ziziphus mauritiana Fruit Pyrus pashia Fruit Gonatanthus pumilus Leaf Ficus semicordata Fruit Zanthoxylum armatum Fruit Castanea sativa Fruit Pyracantha cranulata Fruit 6 11.4.1 Estimation of protein by Lowry’s method Principle The blue colour developed by the reduction of the phosphomolybdicphosphotungstic components in the Folin –ciocalteau reagent by the amino acids tyrosine and tryptophan present in the protein plus the colour developed by the biuret reaction of the protein with the alkaline cupric tartrate were measured in the Lowry’s method. Reagents: i. Folin - Ciocalteau reagent (Reagent A)- A mixture consisting of 100g sodium tungstate (Na2WoO4.2H2O), 25g sodium molybdate (Na2WoO4.2H2O), 700mL water, 50mL of 80% phosphoric acid, and 100mL of concentrated hydrochloric acid was refluxed for 10 hours 66in a 1.5L flask. 150g lithium sulfate, 50mL water and a few drops of bromine water were then added. The mixture was boiled for 15 min without condenser to remove excess bromine. Cool, dilute to 1L and filter. ii. 500 mL of 20% Sodium carbonate in 500 mL 0.1N sodium hydroxide (Reagent B). iii. 0.5% Copper sulphate (CuSO4.5H2O) in 1% potassium sodium tartrate (Reagent C). iv. Alkaline copper solution: Mix 50mL of B and 1mL of C prior to use (Reagent D) v. Protein solution (Stock standard): Fifty mili gram of bovine serum albumin was weighed accurately and dissolve in distilled water and make up to 50mL in a standard flask. vi. Working standard solution: Dilute 10mL of the stock solution to 50mL with distilled water in a standard flask. 1.0mL of this solution contains 200µg protein. Procedure Extraction of protein from sample: Extraction of protien carried out with buffers used for the enzyme assay. 500mg of the sample was grind well with a pestle and mortar in 5-10mL of the buffer. Centrifuge and use the supernatant for protein estimation. Estimation of protein: 1. 0.2, 0.4, 0.6, 0.8 and 1.0mL of the working standard were pipette out into a series of test tubes. 2. Pipette out 0.1 mL and 0.2 ml of the sample extract in two other test tubes. 7 3. Make up the volume to 1.0 ml in all the test tubes with the help of distilled water.A tube with 1.0mL of water without sample serves as the blank. 4. Five mililitre5.0 ml of reagent D was added to each tube including the blank. Mix well and allowed to stand for 10 mins. 5. Then 0.5 mL of reagent A was added. The solution was mixed well and left for incubation at room temperature in the dark. A blue colour was developed after 30 minAbsorbance was noted at 660 nm. A standard graph was drawn and the amount of protein in the sample was calculated with the help of standard curve. 11.4.2 Total Ash Ash value was determined by following the method of AOAC (1990). For this, crucible were kept in a muffle furnace at 450 0 C for 24 h. Then they were transferred from furnace to a desiccator and cooled to room temperature and weighed as quickly as possible to prevent moisture absorption. Two gram dry powder was taken in tared silica crucible and placed in a muffle furnace at 450 0 C for 24 h. Then crucible was transferred to a desiccator and cooled to room temperature, crucible was transferred as quickly as possible to avoid moisture absorption.