Egypt. J. Plant Prot. Res. Inst. (2020), 3 (2): 777 - 785 Egyptian Journal of Plant Protection Research Institute www.ejppri.eg.net Phytochemical and insecticidal evaluation of agro-waste of Lagenaria siceraria (Cucurbitaceae) plant against the cotton leafworm Spodoptera littoralis (Lepidoptera: Noctuidae) Anwaar, M. Abaza Plant Protection Research Institute, Agricultural Research Center, Dokki, Giza, Egypt. ARTICLE INFO Abstract: Article History Lagenaria siceraria (Cucurbitaceae) commonly known as Received: 19/4/2020 bottle gourd cultivated in Africa, Asia and now cultivated in Egypt. Accepted: 28/6/ 2020 After harvesting fruits of L. siceraria, the leaves of the plants are _______________ remained as agro-waste in the field. Preliminary phytochemical Keywords screening of L. siceraria leaves revealed the presence of steroids, Agro-waste, Lagenaria alkaloids, flavonoids, terpenoids, tannins and saponnins. The leaves siceraria, cotton extracted by ethanol and fractionated via solvents of different leafworm, Spodoptera polarities; petroleum ether, ethyl acetate and n-butanol. The toxicity of littoralis, Gas extracts was tested against the cotton leafworm, Spodoptera littoralis Chromatography-Mass (Boisduval) (Lepidoptera: Noctuidae) under laboratory conditions. Spectrometry and The petroleum ether extract exhibited more activity followed by control. ethanol, ethyl acetate and n-butanol. The LC were 0.201, 0.352, 50 0.539 and 0.735%, respectively. The petroleum ether extract was subjected to saponification reaction and its chemical constituents were investigated by Gas Chromatography-Mass Spectrometry. This indicated that the agro-waste of L. siceraria plant can be applied as green insecticide to control the cotton leafworm, S. littoralis. Introduction The Plant kingdom produce a great warmer region of world (Rahman, 2003). variety of phytochemicals (secondary Among all the plants of Cucurbitaceae metabolites) including flavonoids, saponnins, family Lagenaria species is the most popular. tannins, steroids, terpenoids and alkaloids are Lagenaria siceraria (Molina) member of often important for mediating interactions genus Lagenaria that is derived from the between plants and their biotic environment. word lagena, meaning the bottle. L. siceraria They can be models of active defense against known as bottle gourd is a common fruit phytophagous insects and pathogens. vegetable used throughout the India. This Cucurbitaceae family is commonly known as species follows the plant Kingdom : Plantae, gourd, melon and pumpkin family. This Division: Magnoliophyta, Class: family is composed of 118 genera and 825 Magnoliopsida, Order: Cucurbitales, Family: species, which are widely distributed in the Cucurbitaceae, Genus: Lagenaria and Species 777 Abaza, 2020 : L. siceraria L. siceraria large pubescent, compressed with two flat facial ridges annual, prostrate or climbing herb cultivated (Figure, 1) . in Africa, Asia and now cultivated in Egypt, its fruit is a source of vitamin-B complex and choline along with fair source of vitamin-C and β-carotene, amongst any other vegetable known to man till date, it is also good source of minerals and amino acids (Kirtikar and Basu, 2001). The fruits have antimicrobial, cytotoxicity, anticancer and antihepatotoxicity properties (Answar et al., 1984; Desta, 1993; Furukawa et al., 1995 and Shiwaikar and Sreenivasan, 1996). Lagenin, a ribosome inactivating protein (RIP) isolated from the seeds of L. siceraria possesses immune protective, antitumor, anti HIV and anti proliferative properties (Wang and Ng, 2000). After harvesting fruits of L. siceraria, the leaves of the plants are remained as agro- waste in the field. Rahman (2003) described Figure (1): Lagenaria siceraria plant the morphology of this plant species as This study was planned to evaluate follows: Stem: The stem is prostrate or the insecticidal activity of ethanolic extract of climbing in nature with angular, ribbed thick, L. siceraria agro-waste (leaves) and its brittle, softly hairy. Leaves: Leaves are fractions in solvents of different polarities simple with long petiole from 25-30 mm long against the cotton leafworm Spodoptera thick, hallow, densely hairy with two small littoralis (Boisduval) (Lepidoptera: Noctuidae) under laboratory conditions. lateral glands located at the leaf base. Leaf Depending on the insecticidal evaluation, lamina: The lamina of leaf is usually five identify the chemical constituents in the lobed, broad cordate, pubescent with soft bioactive fraction using chromatographic hairs and the tendrils are branched. The techniques. flowers are solitary axillary, pedicellate, Materials and methods unisexual and monoecious. Petals: Petals are 1. Chemicals: mainly five, white or cream colored which The organic solvents of different polarities (petroleum ether, ethyl acetate, opens in the evening. Fruits: Fruits are green ethanol and n-butanol) used in the present in color which turns to yellow on maturity. work were obtained from Edwic Company. Fruits are large densely hairy often 2. Gas Chromatography-Mass cylindrical or flask shaped or globose. They Spectroscopy analysis (GC/MS): are constricted at the middle. Pulp: Pulp is GC-MS analysis was performed on a Varian pale brown in color and the dried fruit has GC interfaced to Finnegan SSQ 7000 Mass selective Detector (SMD) with ICIS V2.0 thick hard hollow structure. Seeds: Seeds are data system for MS identification of the GC embedded in the spongy pulp and 778 Egypt. J. Plant Prot. Res. Inst. (2020), 3 (2): 777 - 785 components. The column used was DB-5 layer was separated in a flask, and the (J&W Scientific, Folosm, CA) cross-linked process was repeated three times to ensure fused silica capillary column (30 m. long, maximum extraction. In the same manner, 0.25mm. internal diameter) coated with poloy ethyl acetate and n-butanol fractions were dimethyl-siloxane (0.5μm. film thickness). obtained, leaving behind a residual aqueous The oven temperature was programmed from fraction. The solvents were evaporated in 50 °С for 3 min., at isothermal, then heating rotary evaporator and the dried fractions were by 7 °С /min. to 250 °С and isothermally for collected. 10 min., at 250 °С. Injector temperature was 6. Preliminary phytochemical screening; 200 °С and the volume injected was 0.5μl. Preliminary phytochemical screening of Transition-line and ion source temperature L. sicerariaA leaves was carried out as the were 250 °С and 150 °С, respectively. The standard prodedures (Sofowara, 1993 and mass spectrometer had a delay of 3 min. to Harborne, 1998). avoid the solvent plead and then scanned 6.1. Alkaloids: The alcoholic extract from m/z 50 to m/z 300. Ionization energy (corresponding to 2.5 g leaves powder) was set at 70 eV. (Agricultural Research was evaporated to dryness and the residue Center, Dokki, Giza). was heated on a boiling water bath with 3. Experimental insect: 2N HCl (2 ml). After cooling the mixture Newly molted 4th instar larvae of the was filtered and the filtrate was treated cotton leafworm S. littoralis were selected for with few drops of Mayer's reagent. this study. The culture of laboratory Mayer's reagent: dissolve 136 g susceptible stain was reared under optimum HgCl2 in 60 ml water and 5 g KI in 10 ml conditions 25±2°C, 75±5% RH. and (16L:8D) water. Combine both solutions and make up light: dark photoperiod in the plant protection with water to 100 ml. add a few drops to an research institute, Dokki, Giza, Egypt. The acidified solution (HCl or diluted H2SO4), a larvae were reared on castor bean leaves yellow to orange precipitate will appear 4. Plant material: indicated a positive test. The agro-waste leaves ofA L. sicerariaA 6.2. Terpenoid: Alcoholic extract was plant were remained after harvesting fruits. L. evaporated and the residue was treated sicerariaAleaves were collected from field of with anhydrous chloroform and filtered. experimental farm in Zagazig Governorate, Conc. sulphuric acid was added to the Egypt and identified by Dr. Reda Mohamed filtrate and shaken. Yellow color was El-Sayed, Faculty of Agriculture, and produced, changed to orange then red. Zagazig University. The collected leaves 6.3. Steroids: Alcoholic extract was were cleaned, left to dry under shade and evaporated and the residue was treated homogenized to fine powder using electric with anhydrous chloroform and filtered. mill. The filtrate was treated with acetic acid 5. Extraction and fractionation and drops of Conc. sulphuric acid were A weighed amount of the powder (250 added carefully on the side of test tube. gm) was soaked in ethanol, the obtained Greenish color appeared (lower layer). extract from filtration was evaporated under 6.4. Flavonoids: A small quantity of the vacuum at 40°C on rotary evaporator to yield extract was dissolved in diluted sodium (45.32 gm). Part of the crude extract (30 gm) hydroxide (5%) and hydrochloric acid was was suspended in distilled water in a added to the mixture. A yellow solution separating funnel, petroleum ether was added that turns colorless on addition of to the funnel which was shaken gently and hydrochloric acid indicated the presence allowed to stand for about 2 hrs, the pet. ether of flavonoids. 779 Abaza, 2020 6.5. Saponins: one gram of the leaves was corrected mortality percentages, the boiled with 10 ml water for few minutes corresponding toxicity lines (LC-P lines) and filtered. The filtrate was vigorously were estimated in addition to determine LC25 shaken. The persistent froth (1 cm height) and LC50 values and their confidence