2.1 Chemicals and sample collection All the chemicals used in this work were of Anal ar grade and purchased from Merck (Germany). Cotton seed of Shimultula was collected from New Market in Dhaka, and seeds of CB-09 and SR-05 were collected from Bangladesh Cotton Development Board in Dhaka, Bangladesh. Healthy disease free and mature fresh fruit samples of cottonseed were collected for analysis. The seeds external part was removed, rinsed thoroughly with running water and dried in oven at 105°C till constant weight. The dry seeds were grounded into fine particles using blender. All tests were carried out three times using dry sample and the analytical results presented are the average values.
2.2 Extraction of oil from cottonseeds For the extraction of oil from cottonseeds, petroleum ether (b.p. 40−60°C) was used to extract oil from 100 g of powder using a soxhlet apparatus for 72hrs. In order to remove solvent, oil extract was filtered followed by vacuum distillation. The color of the extracted oil was chocolate maroon. In final stage, activated charcoal was used to filter the extracted oil which gave the light golden color of the oil. The oil content was calculated and preserved in a refrigerator for further analysis.
2.3 Proximate analysis The proximate analysis of cottonseed was performed using authorized methods developed by Association of Official Analytical Chemistry (AOAC) [AOAC, 1990].
2.4 Determination of the physicochemical characteristics of the oil The physicochemical properties of the extracted oil were investigated as per cited standard methods [Kirk et al., 1991; Anonymous, 2004].
2.5 Chemical analysis of fatty acids Gas Chromatography technique (GC-FID) was used for the analysis of fatty acids (Shimadzu GC-2025, Detector: Flame ionization detector (FID), Column: capillary type (Capillary HP 5), packing material: 10% diethylene glycol succinate on 100-120 mesh, carrier gas: N2 with flow rate 1 mL/min). The fatty acid methyl esters (FAMEs) were prepared by complete esterification of oil using BF3−MeOH complex (Mariod, 2011; Kirk et al., 1991). Standard FAMEs were collected from Sigma chemical Co, St Louis, Mo, USA (Fatty acid methyl ester containing 11 standards such as caprilic acid, lauric acid, myristic acid, palmitic acid, palmitoleic acid, linoleic acid, oleic acid, stearic acid, arachidonic acid, behenic acid, lignoceric acid and arachidic acid) were used for the identification and quantification of the peaks. Each FAME in extracts was identified by comparing retention times.
2.6 Analysis of Mineral composition cottonseeds Estimation of mineral content such as Mg, P, Zn, S, K, Ca, Cr, Mn, Fe, Ni, B, Pb, Cd etc. were carried out in the fresh seed powder on dry weight basis. Elemental analysis was conducted by using Atomic Absorption Spectroscopy (AAS) technique (Model: SpectrAA 55B, origin: Australia. Instrument type: wave length dispersive AAS (WDXRF) with SC and PC type detector, light source: X-ray tube anode Rh tube, fully automated). The values of macronutrients are expressed in percentage (g/100g) and micronutrients were expressed as mgKg-1on dry weight basis.