Variability in permeability and integrity of cell membrane and depletion of food reserves in neem (Azadirachta indica) seeds from trees of different age classes

Abstract

We quantified cell membrane permeability (electrical conductivity-EC, water soluble sugar-WSS, and amino acids-AA) and integrity (phospholipids, α-tocopherol and lipid peroxidation) along with food reserve deterioration (total proteins, total sugar, and total starch) of neem seeds collected from various mother tree age classes and stored for 65 days in airtight plastic containers at ambient room temperature (35±5°C). Results show that the activities were higher in fresh seeds (EC 267.56–2950.01 μS/g, WSS 19.96–19.48 mg/g and AA 5.40–5.35 mg/g) and declined with increasing duration of storage period (EC 153.37–195.17 μS/g, WSS 3.13–4.17 mg/g and AA 4.29–4.49 mg/g after 35 days and EC 144.02–161.56 μS/g, WSS 2.06–2.40 mg/g and AA 3.98–4.27 mg/g after 65 days of storage). Phospholipids and α-tocopherol were higher in fresh seed (0.073–0.093 OD at 710 nm and 0.080–0.105 OD, respectively) and declined as storage duration increased (0.033–0.042 OD at 710 nm and 00.0010–0.0020 OD, respectively). Dead seeds showed reduced amounts of phospholipids and minimum activity of α-tocopherol (antioxidants). The level of MDA was lower in fresh seeds (0.0066–0.0087 OD at 600–535 nm) and increased as storage duration increased (0.0248–0.0268 OD after 65 days of storage). The higher amount of MDA indicated that seeds died due to rancidity of the oil inside the seed. Neem seed cake was assessed for deterioration of food reserves (total proteins, total sugar, and total starch), concentrations of which were higher in fresh seed and declined as storage duration increased. Germination was higher in fresh seeds and after 65 days, no germination was received perhaps due to deterioration of biochemicals in seeds. Patterns of seed deterioration were similar across all seed lots.

Keywords

phospholipids / α-Tocopherol / Lipid peroxidation / Azadirachta indica

References

[1] Abdul-Baki AA, Anderson JD. Viability and leaching of sugars from germinating barley. Crop Science, 1970, 10: 31-34.

[2] Abdul-Baki AA, Anderson JD. Kozlowski TT. Physiological and biochemical deterioration of seeds. Seed biology. 1972, New York: Academic Press, 283 315

[3] Agrawal PK, Dadlani M. Agrawal PK, Dadlani M. Membrane permeability: A measure of seed quality. Techniques in seed science and technology. 1992, New Delhi: South Asian Publisher, International Book Company Absecon Highlands, 127 133

[4] Agrawal PK. Germination, fat acidity and leaching of sugars from five cultivars of paddy rice (Oryza sativa) seeds during storage. Seed Science and Technology, 1977, 5: 489-498.

[5] Anderson JD. Enzymes and their role in wheat technology. 1946, New York: Inter science Publishers Inc., 371.

[6] Anon. Progress report of ICFRE Scheme. All India coordinated research project for investigation on agricultural byproducts and industrial waste materials for evolving economic rations for livestock. 1978, Jabalpur, India: Jawahar Lal Nehru Krishi Vishva Vidyapith

[7] AOAC Official Methods for Analysis. Association of official analytical chemists, 1990 15th Ed. USA: Incorporation Virginia, 951 986

[8] Balkrishnan B, Nair MC. Observation on the post infectional changes of rice plants infected by the sheath rot fungus, Acrocylindrium oryzae saw. Agriculture Research Journal of Kerala, 1983, 21: 73-74.

[9] Barber AA, Bernheim F. Strehler BL. Lipid peroxidation: its measurement, occurrence, and significance in animal tissues. Advanced Genetic Research. 1967, New York: Academy Press, 355 403

[10] Bewley JD, Black M. Physiology and Biochemistry of seeds (Vol. 2). 1982, New York: Springer-Verlag, 1 58

[11] Bewley JD. McDonald MB, Nelson CJ. Membrane changes in seeds as related to germination and the perturbations resulting from deterioration in storage. Physiology of seed deterioration. 1986, Madison, WI: Crop Science Society of America, 27 46

[12] Bonner FT. Estimating seed quality of southern Pines by leachate conductivity, 1991 1 4

[13] Bradnock WR, Matthews S. Assessing field emergence potential of wrinkle seeds peas. Horticulture Research, 1970, 10: 50-58.

[14] Cherry JP, Mayre RV, Ory RL. Protein content and enzymes in rare seeds of Arachis hypogae L. Physiological Plant Pathology, 1974, 41: 425-439.

[15] Ching TM, Scoolcraft I. Physiological and chemical differences in aged seeds. Crop Science, 1968, 8: 407-409.

[16] Colowic SP, Kaplan WC. Method in Enzymology (Vol. III). 1957, New York: Academic Press, 468.

[17] Dadlani M, Agrawal PK. Mechanism of soybean seed deterioration. Plant Physiology and Biochemistry, 1983, 10: 23-30.

[18] Djavanshir K, Pourbeik H. Germination value -a new formula. Silvae Genetica, 1976, 25: 79-83.

[19] Dubois M, Gilles KA, Hamilton JK, Robers PA, Smith F. Colorimetric method for determination of sugar and related substrates. Annals of Chemistry, 1956, 28: 350-356.

[20] Ezumah BS. Germination of storage of neem (Azadirachta indica A. Juss.). Seed Science and Technology, 1986, 14: 593-600.

[21] Fick GL, Hibbard RP. A method for determining seed viability by electrical conductivity measurements. Michigan Academy of Science arts and Letters, 1952, 5: 95-103.

[22] Fielding JL, Goldworthy A. Tocopherol levels and ageing in wheat grains. Annals of Botany, 1980, 46: 453-456.

[23] Ghose B, Chaudhari MM. Ribonucleic and breakdown and loss of protein synthetic capacity with loss of viability of rice embryo. Seed Science and Technology, 1984, 12: 669-677.

[24] Halder S, Kole S, Gupta K. On the mechanism of sunflower seed deterioration under two different type of accelerated ageing. Seed Science and Technology, 1983, 11: 331-339.

[25] Helmer JD, Delouche JC, Leinhard M. Some indices of vigor and deterioration in seeds of crimson clover. Proceeding Association Official Seed Analysis, 1962, 52: 154-160.

[26] Heydecker W. Roberts EH. Vigor. Viability of seeds. 1972, Syracuse, N.Y.: Syracuse University Press, 209 252

[27] Hsu FH, Lin JB, Chang SR. Effects of water logging on seed germination, electric conductivity of seed leakage and developments of hypocotyls and radicle in sudangrass. Botanical Bulletin of Academia Sinica, 2000, 41: 267-273.

[28] Kaloyereas SA. Rancidity as a factor in the loss of viability of pine and other seeds. Journal of the American Oil Chemists Society, 1958, 35: 176-179.

[29] Koostra PT, Harrington JF. Biochemical effects of age on membrane lipids of Cucumis sativus L. seed. Proceedings of the International Seed Testing Association, 1969, 34: 329-340.

[30] Lowery OH, Rosebroagh NJ, Farr AL, Randall RJ. Protein measurement with the folin phenol reagent. Journal of Biological Chemistry, 1951, 193: 265-275.

[31] Maccomb CL, Winstead NN. Changes in sugar and aminoacids of cucumber fruits infected with Pythium aphanidermatum. Phytopathology, 1964, 54: 233-234.

[32] Mani G. Seed Technology studies in Acacia nilotica (Linn.) and Acacia leucocephloea. 1997, Coimbatore-641003: Department of Seed Technology, Tamil Nadu Agricultural University

[33] Matthews S, Bradnock WT. The detection of seed samples of wrinkled-seeded peas (Pisum sativum L.) of potentially low planting value. Proceedings of the International Seed Testing Association, 1967, 32: 555-563.

[34] Matthews S, Whitbread R. Factors affecting pre-emergence mortality in peas. I. An association between seed exudates and the incidence of pre-emergence mortality in wrinkle-seeded peas. Plant Pathology, 1967, 17: 11-17.

[35] Mukhtar ND, Laidman DL. Mineral ion transport in the embryos of germinating wheat (Triticum aestivum). Journal of Experimental Botany, 1982, 33: 643-655.

[36] Nagaveni HC, Ananthapadmanabha HS, Rai SN. A note on the extension of viability of Azadirachta indica A. Juss. Myforest, 1987, 23 245.

[37] Nautiyal AR, Purohit AN. Seed viability in Sal I. Physiological and biochemical aspects of seed development in Shorea robusta. Seed Science and Technology, 1985, 13: 59-68.

[38] Nema DK. Changes in sugar of apple due to Alternaria alternata infection. Indian Phytopath, 1983, 36: 624-629.

[39] Niehaus WJ Jr.. A proposed role of superoxide anion as a biological nucleophile in the desertification of phospholipid. Bioorganic Chemistry, 1978, 7: 77-84.

[40] Parmar BS, Katkar CM. Randhawa NS, Parmar BS. Commercialization. Neem-1993, Publisher, Society of Pesticides Science India and New Age International (P) Ltd, 1996 318 332

[41] Pearce RS, Abdel Samad IM. Change in fatty acid content of polar lipids during ageing of seeds of peanut (Arachis hypogiea L.). Journal of Experimental Botany, 1980, 31: 1283-1290.

[42] Pearson D. 1970. The Chemical Analysis of Foods. New York: Chemical Publishing Co., p.535.

[43] Pehap A. Selected papers in gerontology of some forest seeds. Research Notes, Skosgshogskolan. 1973, Stockholm: Royal College of Forestry

[44] Perry CA. Handbook of Vigor Test Methods, 1987 2nd ed. Zurich: ISTA

[45] Pesis E, Timothy JN. Viability, vigor and electrolytic leakage of muskmelon seeds subjected to accelerated ageing. Horticulture Science, 1983, 18: 242-244.

[46] Powell AA, Mathews S. Deteriorative changes in pea seeds (Pisum sativum L.) stored in humid or dry conditions. Journal of Experimental Botany, 1977, 28: 225-234.

[47] Powell AD, Dulson J, Bewley JD. Changes in germination and reparatory potential of embryos of Grand Rapid lettuce seeds during long term imbibed storage, and related changes in the endosperm. Planta, 1984, 162: 40-45.

[48] Powell AD, Leung DWM, Bewley JD. Long term storage of dormant Grand Papids lettuce seeds in the imbibed state: physiological and metabolic changes. Planta, 1983, 159: 182-188.

[49] Prasad BK. Free sugar and amino acids contents of coriander infected with stem gall disease. Indian Phytopathology, 1983, 36: 150-151.

[50] Presley DA, Mcbride MB, Leopold AC. Tocopherol and organic free radical levels in soybean seeds during natural and accelerated aging. Plant Physiology, 1980, 66: 715-719.

[51] Priestley DA, Leopold AC. Absence of lipid oxidation during accelerated aging of soybean seeds. Plant Physiology, 1979, 63: 726-729.

[52] Priestley DA. Seed ageing: Implication of seed storage and persistence in the soil. 1986, Ithaca: Cornell University Press, 304.

[53] Pukacka S, Czubak A. The effect of desiccation on viability and membrane lipid composition of Acer pseudoplatanus seeds. Acta Societatis Botanicorum Poloniae, 1998, 67: 249-252.

[54] Pukacka S, Ratajczak E. Age related biochemical changes during storage of beech (Fagus sylvatica L.) seeds. Seeds Science Research, 2007, 17: 45-53.

[55] Pukacka S. Changes in membrane fatty acids composition during desiccation of seeds of Acer saccharinum (silver maple). Seed Science and Technology, 1998, 26: 535-540.

[56] Raheja RS, Kaur C, Singh A, Bhatia IS. New colorimetric method for the quantitative estimation of phospholipid without acid digestion. Journal Lipid Research, 1973, 14: 695-697.

[57] Rawat MMS, Thapliyal RC. Endogenous rhythm in seed germination of Dendrocalamus strictus. Seed Science and Technology, 2003, 31: 21-27.

[58] Roberts BE, Payne PI, Osborne DJ. Loss of activity of protein synthesizing systems in vitro associated with a loss of viability. Biochemistry Journal, 1973, 131: 275-286.

[59] Simon EW, Raja Harun RM. Leakage during seed imbibition. Journal of Experimental Botany, 1972, 23: 1076-1085.

[60] Singh P, Singh O, Chandra V. Estimating seed quality in hard seeded leguminous trees by accelerated aging and leachate conductivity. Indian Forester, 1996, 122: 415-418.

[61] Stewart RRC, Bewlay JD. Lipid peroxidation associated aging of soybean axes. Plant Physiology, 1980, 65: 245-248.

[62] Suszka B. Cold storage of already after ripened beech (F. silvatica) seeds. Arboretum Kornickie, 1975, 20: 299-315.

[63] Takayanagi K, Murakami K. Rapid germinability test with exudates from seeds. Nature, 1968, 218: 493-494.

[64] Tammela P, Hopia A, Hiltunen R, Vuorela H, Nygren M. Ageing in Pinus sylvestris L. seeds: Changes in viability and lipids. Biochemistry Society Trans, 2000, 28: 878-879.

[65] Tappel AL. Free radical peroxidation damage and its inhibition by Vitamin E and selenium. Fed Proc Fed Am Socs Exp Biol, 1965, 24: 73-78.

[66] Tripathi YC, Gupta PK, Rathore M. Biodeterioratve effect on chemical composition of neem seeds. Bulletin of Pure and Applied Science, 1996, 15B: 121-124.

[67] Umarani R. Studies on the physiological and biochemical basis of seed germination and deterioration in Casuarina equisetifilia. 1999, Coimbatore-641003: Tamil Nadu Agricultural University

[68] Verghees B, Naithani SC. Physiological and biochemical changes associated with ageing in neem seed. IUFRO Symposium on innovation in forest tree seed science and nursery technology. 1997, Raipur-492010, India: Pt. Ravishankar Shukla University, 157 158

[69] Varghese B, Naithani SC. Desiccation induced changes in lipid peroxidation, superoxide level and antioxidant enzymes activity in neem (Azadirachta indica A. Juss.) seeds. Acta Physiol Plant, 2002, 4(1): 79-87.

[70] Vijayaragavan A, Vanangamudi K. Effect of seed source and period of storage on germination and biochemical activity of Albizia lebbeck (L) Benth. Seeds. My Forest, 2003, 39: 341-351.

[71] Villiers TA. Seed aging: Chromosome stability and extended viability of seeds stored fully imbibed. Plant Physiology, 1974, 53: 875-878.

[72] Walker BL, Singer SJ. Effect of processing on the tocopherol content of rapeseed oil. Canadian Institute of Food Science and Technology Journal, 1975, 8: 179-180.

[73] Wilson DO, Mcdonald JrMB. The lipid peroxidation model of seed ageing. Seed Science and Technology, 1986, 14: 269-300.

[74] Yaklich RW, Abdul-Baki AA. Variability in metabolism of individual axes of soybean seeds and its relationship to vigor. Crop Science, 1975, 15: 424-426.

[75] Yamauchi R, Matsushita S. Quantitative changes in tocopherols and their intracellular distribution in cotyledons during soybean germination. Nippon Nogeikagakn Kaishi, 1976, 50: 525-528.