BOTANICAL OILS OF FENUGREEK, PUMPKIN AND SWEET ALMOND: GAS CHROMATOGRAPHY- MASS SPECTROMETRY (GC-Ms) ANALYSIS, ITS EFFECTS ON MULBERRY SILKWORM, BOMBYX Mori L.

Abstract

The present investigations were carried out during two spring seasons, 2019-2020 in the silkworm laboratory of Sericulture Department of Plant Protection Research Institute, Sharkia Branch to investigate the effect of fenugreek, pumpkin and sweet almond oils on silk production of mulberry silkworm Bombyx mori L. The obtained results cleared that all of these plants oils, fenugreek; pumpkin and sweet almond oils composed of many methylated fatty acids fraction which have many effects of fresh cocoon weight, shell cocoon weight, silk ratio, silk filament weight and size, whereas the fenugreek oil consisted of forty-eight compounds (Hexadecanoic acid, methyl ester, 9-Octadecenoic acid (Z)-, methyl ester, 9,12-Octadecadienoic acid (Z,Z)-, methyl ester and Hexadecanoic acid, methyl ester etc.....), pumpkin consisted of seventy-two ( 9,12Octadecadienoic acid (Z,Z)-, methyl ester, Hexa decanoic acid, methyl ester, Methyl stearate, Dodecanoic acid, methyl ester and Tridecanoic acid, 12-methyl-, methyl ester etc.....)and sweet almond oil consisted of sixteen compounds (Hexadecanoic acid, methyl ester, 9,12Octadecadienoic acid (Z,Z)-, methyl ester, 9-Octadecenoic acid (Z)-, methyl ester, Methyl stearate and Hexanal dimethyl acetal etc....). Fenugreek, pumpkin and sweet almond oils improved cocoon indices and silk filament characters as a profitable supplementary diet for silk worm Bombyx mori L. Conclusively, the fenugreek, pumpkin gave the highest mean of silk filament length. Fenugreek, sweet almond and pumpkin oils improved cocoon indices and silk filament characters as a profitable supplementary diet for silk worm Bombyx mori L. keywords: Fenugreek, Sweet almond, pumpkin oils, Mulberry silkworm Larvae, Bombyx Mori L. 720 WALAA SALLAM et al. INTRODUCT: The silk industry depends on the growth of silkworm that depends on body weight increase and the biochemical components accumulation of food like protein, amino acids and enzymes (Eman and Marwa 2017). The plants oils could be used for nutrition silkworm to improve silkworm development and silk production, they are making as antimicrobial activity with no side effects on it (Eman and Marwa 2017, Tabassum and Vidyasagar 2013). They have a nutrition profile as a good source of (vitamins E, niacin, riboflavin, etc.), minerals (potassium, phosphorus, magnesium, calcium, iron, zinc, manganese, selenium, copper, etc.), macronutrients (proteins, carbohydrates, fibers, etc.) and fatty acid (saturated, monosaturated and polyunsaturated). The chemical and nutritional composition have been shown to be significantly decrease the risk of diseases, antioxidation and anti-inflammatory action. Therefore, the aim of this study to evaluate botanical oils of fenugreek, pumpkin and sweet almond: gas chromatographymass spectrometry (gc-ms) analysis, its effects on mulberry silkworm, Bombyx mori L. MATERIALS AND METHODS The present investigations were carried out during two spring seasons, 2019 and 2020 in silkworm laboratory of Sericulture Department of Plant Protection Research Institute, Sharkia Branch. The following experiments were carried out: 1Feeding technique: The tested materials were used solely at three concentrations (1, 2 and 3 %). Three replicates of 30 silkworm larvae for each concentration were fed, throughout 4th &5th instars on mulberry leaves sprayed on the tested concentrations of each material then left to dry under laboratory conditions. Three replicates were sprayed in distilled water as control, larvae of each replicate were reared on a plastic tray (100 × 70 × 15 cm) under at laboratory conditions 27±2oC and 95±5 % RH for the first three instar (1-3), while it was changed for the last two (fourth and fifth instars) to 24±2 oC and 75±5 % RH. All rearing trays, racks and tools, as well as the rearing rooms were sterilized with formalin (5%) one week before the beginning of the experiment. The tested larvae were fed daily at 8 am, 12 noon, 4 and 8 pm. Five full grown larvae/ replicate were weighted, individually then the two silk glands of each larva were dissected out on a filter paper then weighed on a digital balance. Mature larvae were handly transferred to carton paper (that used wrapping table eggs) for mounting process (Zannoon and Shadia, 1994). The cocoons were harvested 7 days later. Five freshes of the resulted cocoons of each replicate were taken for measuring the cocoon indices (fresh cocoon J. Product. & Dev., 26(4),2021 721 weight, cocoon shell weight and silk ratio), whereas, half of rest cocoons were dried in an oven (oven temperature was raised gradually until it reached 80°C for 6 hour) to study filament characters (filament length, filament weight and filament size). Another half of the cocoons were used for the biological studies. After emergence, each couple was impaired in paper saccule for copulation and oviposition. 2Spraying leaf technique: In this trail, the leaves spraying technique was used to evaluate the effect of doses of some concentrations for the tested compounds on the 4 th and 5 th instars larvae. The sprayed leaves were left to dry and provide to larvae (30 larvae for each concentration) in treatments and control. The biology, physiology and productivity measures were investigated. 3Measurements: The technological parameters were measured in both experiments as follows: 4.1Technological parameters: 4.1.1. Cocoon indices: The following parameters were measured: AFresh cocoon weight (g). BCocoon shell weight (g). CCocoon shell ratio = × 100 (Krishnaswami, 1978). 4.1.2. Reelable silk filament parameters: The weight (g) and length (m) of reeled silk filament were measured and recorded. The size of the reeled filament (denier) was estimated according to (Krishnaswami, 1978) formula: The size of reeled filament = × 9000 Data obtained were statistically analyzed according to Snedecor and Cochran (1982) methods using software COSTAT program. 4.2. BOTANICAL STUDIES: 4.2.1. Methylation: The H2SO4 derivatization method has also been widely used for the analysis of fatty acids in biological samples.H2SO4 -methanol 2% (v/v) was added into a vial containing previously weighed 10 mg of fat. The vial was heated at 80 C with occasional shaking. Afterwards, 0.25 mL of the neutralized aqueous solution (sodium hydroxide 1 M) was added, and it was smoothly shaken. Before the analysis of the n-hexane phase, the sample was allowed to rest for 5 min (Masood et al., 2005). The reaction procedure is similar to that 722 WALAA SALLAM et al. of other derivatization methods. Because H2SO4 is a strong oxidizing agent, this method is not recommended for PUFA analysis (Kuksis, 1994; Michal Ski and Łyko, 2010). 4.2.2. Gas chromatography–mass spectrometry (GC-MS) analysis: The chemical composition of plant oils was performed using Trace GC1310-ISQ mass spectrometer (Thermo Scientific, Austin, TX, USA) with a direct capillary column TG–5MS (30 m x 0.25 mm x 0.25 μm film thickness). The column oven temperature was initially held at 50 C and then increased by 5°C /min to 230°C hold for 2 min. increased to the final temperature 290°C by 30°C /min and hold for 2 min. The injector and MS transfer line temperatures were kept at 250, 260°C respectively; Helium was used as a carrier gas at a constant flow rate of 1 ml/min. The solvent delay was 3 min and diluted samples of 1 μl were injected automatically using Autosampler AS1300 coupled with GC in the split mode. EI mass spectra were collected at 70 eV ionization voltages over the range of m/z 40–1000 in full scan mode. The ion source temperature was set at 200 °C. The components were identified by comparison of their retention times and mass spectra with those of WILEY 09 and NIST 11 mass spectral database. RESULTS AND DISCUSSION: Part 1: Effect of some plant oils on the technology, and silk production of