Development of an RP-UHPLC-PDA method for quantification of free gossypol in cottonseed cake and fungal-treated cottonseed cake

Abstract

Cottonseed cake biomass, which is a residue of oil extraction, is potentially appropriate for use as animal feed, given the high mineral, fibre and protein content. The presence of free gossypol, however, a toxic pigment in the glands of the cotton plant, limits use of this biomass for monogastric livestock. A promising method to detoxify cottonseed cake relies on fermentation by fungi, which can eliminate up to 100% of gossypol. In order to quantify trace levels of free gossypol in different cotton materials, including cottonseed cake treated with macrofungi, a simple and rapid chromatographic detection method was developed and validated. Under optimized conditions, extraction was performed using 70% acetone. The extract was then analysed by Ultra High-Performance Liquid Chromatography (UHPLC), with gradient elution on a C18 reverse phase column KINETEX® (100 x 2.10 mm, 2.6 μm). Methanol-0.1% TFA aqueous solution was employed as mobile phase and PDA detection conducted at 254 nm. The optimized method was validated by analysis of specificity, linearity and range, limit of detection, limit of quantification, precision and accuracy. Detection and quantification limits were observed at 0.2 and 0.5 μg/mL, respectively. With good reproducibility, with precision (RSD)<10% and recovery greater than 94%, the developed assay was appropriate for quantification of low quantities of free gossypol. The validated method was successfully applied to determine trace levels of free gossypol cottonseed treated with a macrofungus.

Evaluation of Bio-detoxification of Jatropha curcas Seed Cake and Cottonseed Cake by Basidiomycetes: Nutritional and Antioxidant Effects

“…Besides visual measurements in centimeters, another indicator of fungi colonization is the presence of ergosterol [44,26]. This molecule, vitamin D 2 precursor, is sterol present in cellular membranes of fungi and commonly used as an indicator of fungal growth [45,46,25]. Some authors have also reported higher levels of ergosterol in basidiomycetes species [47,48,46], showing a positive correlation between fungi growth and ergosterol content [47,48], which corroborates the use of this molecule as a fungal biomass growth indicator.…”

“…Ergosterol, a primary sterol present in cell membranes of lamentous fungi is widely used to estimate fungal biomass [25]. Ergosterol quanti cation was performed in triplicate, applying a method adapted from a published protocol [26].…”

Biodegradation of Gossypol by Aspergillus terreus-YJ01

“…Then, 1.0 mL of acetonitrile was added to the bottom precipitation, and the gossypol in the precipitation was fully dissolved in acetonitrile using ultrasonic oscillation. The supernatant was diluted and used to determine the concentration of gossypol busing high-performance liquid chromatography (HPLC) [ 32 , 33 ]. The data are presented as the average values derived from three measurements, and their relative standard deviations were less than 10%.…”

Cottonseed Meal Protein Isolate as a New Source of Alternative Proteins: A Proteomics Perspective

“…The air-dried free and total gossypols were dissolved in 70% acetone and analysed using the Dionex Ultimate TM 3000 UHPLC Systems (Thermo Fisher Scientific, Waltham, MA, USA). The analysis method of Conceicao et al was performed with minor modifications [ 57 ]. Gradient elution was performed using a mobile phase beginning with 40% aqueous (TFA 0.1%, v / v ) and 60% organic (100% methanol) solvent on a KINETEX ® EVO C18 column (150 × 3.0 mm, 2.6 μm) (Phenomenex, Torrance, CA, USA) at 35 °C.…”