Michael K. Dowd

Effect of Racemic and (+)- and (−)-Gossypol on the Survival and Development of Helicoverpa zea Larvae

Gossypol is a sesquiterpene that occurs naturally in seed and other parts of the cotton plant. Because of restricted rotation around the binaphthyl bond, it occurs naturally as enantiomeric mixtures with (+)-gossypol to (−)-gossypol ratios that vary between 97:3 and 31:69. Commercial cotton varieties (Gossypium hirsutum) normally exhibit an approximate 3:2 ratio. (+)-Gossypol is significantly less toxic than (−)-gossypol to nonruminant animals; thus, cottonseed containing high levels of (+)-gossypol might be safely fed to nonruminants. Gossypol, however, is an important component in the cotton plants defense against insect herbivores, but it is not known how cotton plants that exhibit high levels of (+)-gossypol in the foliage might be affected by insect herbivory. To address this question, 1-d-old Helicoverpa zea larvae were fed diets with 0.16, 0.20, and 0.24% racemic, (+)-, and (−)-gossypol. Larval pupal weights, days-to-pupation, and survival were adversely affected by all gossypol diets compared with the control diet. Statistical differences were determined by comparing the compounds among themselves at the three levels and between the three compounds at the same level. When the compounds were compared among themselves, no large differences were observed in pupal weights or in days-to-pupation among any of the diets. Among the three compounds, at the 0.16% level, the diet containing racemic gossypol was the most effective at reducing survival. At the 0.20 and 0.24% levels of racemic (+)- and (−)-gossypol, survival was not statistically different. The overall results indicate that (+)-gossypol is as inhibitory to H. zea larvae as racemic or (−)-gossypol, and thus, cotton plants containing predominantly the (+)-enantiomer in foliage may maintain significant defense against insect herbivory.

Induction of hairy root cultures from Gossypium hirsutum and Gossypium barbadense to produce gossypol and related compounds

Hairy root cultures were induced by inoculating cotyledonary leaves and hypocotyl segments from two cotton species, Gossypium hirsutum and Gossypium barbadense, with Rhizobium rhizogenes 15834. For both species, more hairy roots formed on inoculation sites on cotyledonary leaves than on hypocotyls. The addition of sucrose to basal Murashige–Skoog media increased the frequency of hairy root formation, whereas the addition of naphthalene acetic acid (0.54xa0μM) did not. After transfer to a liquid culture, hairy root growth was very rapid. After 3xa0wk in liquid culture, both cotton species produced gossypol, a di-sesquiterpene secondary metabolite with known anticancer activity, and two related methylated derivatives. Most (60–95%) gossypol produced by cultures was retained within the hairy root tissues, but some was found in the media. The average gossypol level observed among 96 different cultures was 15xa0mg/g of dry culture mass; however, some cultures produced >40xa0mg/g of dry culture mass. Variation in gossypol levels was greater for cultures arising from different transformation events than for multiple subclones of a single transformant. The high level of gossypol production attained by most of these cultures suggests that they will be valuable for studying the biochemical and molecular aspects of gossypol biosynthesis, capable of producing large amounts of gossypol and related compounds, and useful for generating modified forms of gossypol (e.g., radio-labeled gossypol) for understanding bioactivity mechanisms.

Effect of Racemic, (+)- and (−)-Gossypol on Survival and Development of Heliothis virescens Larvae

Gossypol is a constituent of the lysigenous foliar glands of cotton plants and is also found in glands in cottonseed. Gossypol exists as enantiomers because of restricted rotation around the binaphthyl bond. The biological activities of the enantiomers differ. For example, (+)-gossypol can be fed safely to nonruminants such as chickens, but (−)-gossypol cannot. Most commercial cottonseed contain a (+)- to (−)-gossypol ratio of ≈3:2. Conventional breeding techniques can be used to develop cottonseed that contains >95% (+)-gossypol. Notably, gossypol protects the plant from insect herbivores. Herein, we report the effect of various forms of gossypol on Heliothis virescens (Fabricius) larvae. Three levels (0.16, 0.24, and 0.32%) of racemic, (+)-, and (−)-gossypol were added to artificial rearing diets and were fed to H. virescens larvae. All 0.24 and 0.32% gossypol diets significantly lengthened days-to-pupation and decreased pupal weight compared with the control. Percent survival was significantly less for larvae reared on diets containing 0.24% of all three forms of gossypol as compared with the control diet. (+)-Gossypol was superior or equivalent to racemic gossypol as measured by the three parameters studied. Higher concentrations of all gossypol forms were required to reduce survival and pupal weights and increase days-to-pupation for larvae of H. virescens larvae compared with the concentration needed to affect larvae of Helicoverpa zea (Boddie), which was studied previously. These results indicate that current efforts to breed cotton lines containing mostly (+)-gossypol in seed should not significantly impair the plant’s natural defenses against insects.

Alternative Hydrocarbon Solvents for Cottonseed Extraction: Plant Trials

Hexane has been used for decades to extract oil from cottonseed and is still the solvent of choice for the edible-oil industry. Due to increased regulations as a result of the 1990 Clean Air Act and potential health risks, the edible-oil extraction industry urgently needs an alternate hydrocarbon solvent to replace hexane. Based on laboratory-scale extraction tests, two hydrocarbon solvents, heptane and isohexane, were recommended as potential replacements for hexane. A cottonseed processing mill with a 270 MT/day (300 tons/day) capacity agreed to test both solvents with their expander-solvent process. Extraction efficiencies of isohexane and heptane, judged by extraction time and residual oil in meal, refined and bleached color of miscella refined oil, and solvent loss, were comparable to that of hexane. However, fewer problems were encountered with the lower-boiling isohexane than with the higher-boiling heptane. With isohexane, the daily throughput increased more than 20%, and natural gas consumption decreased more than 40% as compared to hexane.

Wood adhesive properties of cottonseed protein with denaturant additives

Abstract Most commercial wood adhesive use either formaldehyde-based resins or polyurethanes, both of which include potentially toxic chemicals in their formulations. As a result, proteins are being considered as greener and more sustainable wood adhesives. While most of the protein adhesive studies focus on soy proteins, there is also interest in exploring alternatives. In this work, testing of the adhesive performance of cottonseed protein isolate was undertaken in the presence of protein denaturants, i.e. guanidine hydrochloride (GuHCl), sodium dodecyl sulfonate (SDS), urea, and alkali. For comparison, soy protein isolate was also included in the study. At optimal dosage levels, the dry adhesive strength of cottonseed protein isolate could be enhanced by 38, 25, or 47% with SDS, GuHCl, or urea, respectively. The dry adhesive strength and hot water resistance of cottonseed protein isolate was generally superior to that of soy protein isolate, with or without the denaturants. Thus, the combination of cottonseed protein with an optimal concentration of a denaturant may be a potentially promising polymeric system for use as wood adhesives.

Naturally occurring high oleic acid cottonseed oil: identification and functional analysis of a mutant allele of Gossypium barbadense fatty acid desaturase-2

AbstractMain conclusionSome naturally occurring cotton accessions contain commercially attractive seed oil fatty acid profiles. The likely causal factor for a high-oleate trait in pima cotton (Gossypium barbadense) accession GB-713 is described here.n Vegetable oils are broadly used in the manufacture of many human and animal nutritional products, and in various industrial applications. Along with other well-known edible plant oils from soybean, corn, and canola, cottonseed oil is a valuable commodity. Cottonseed oil is a co-product derived from the processing of cottonseed fiber. In the past, it was used extensively in a variety of food applications. However, cottonseed oil has lost market share in recent years due to less than optimal ratios of the constituent fatty acids found in either traditional or partially hydrogenated oil. Increased awareness of the negative health consequences of dietary trans-fats, along with the public wariness associated with genetically modified organisms has created high demand for naturally occurring oil with high monounsaturate/polyunsaturate ratios. Here, we report the discovery of multiple exotic accessions of pima cotton that contain elevated seed oil oleate content. The genome of one such accession was sequenced, and a mutant candidate fatty acid desaturase-2 (FAD2-1D) gene was identified. The mutant protein produced significantly less linoleic acid in infiltrated Arabidopsis leaf assays, compared to a repaired version of the same enzyme. Identification of this gene provides a valuable resource. Development of markers associated with this mutant locus will be very useful in efforts to breed the high-oleate trait into agronomic fiber accessions of upland cotton.

Fundamental research on spiking, recovery and understanding seed coat nep counts in AFIS analysis of pre-opened cotton:

Understanding seed coat fragment (SCF) spiking results in advanced fiber information systems (AFIS) analysis of seed coat neps (SCN) in ginned cottons is confounded by the opening of entangled fibers in the instrument’s fiber individualizer. This may influence seed coat tissue fragmentation and recovery since a high degree of machine-fiber interaction is required to individualize entities for sensing. In this paper, slivers were pre-opened in the AFIS followed by manual cleaning, spiking, and AFIS analysis. A protocol was developed to spike the pre-opened slivers. The percent recovery of the spiked entity was dependent on the species and cultivar of the cotton used to prepare the slivers. The lowest recovery was with Pima fibers. Also, the recovery improved with the increase in length of the fibers biologically attached to the SCF surface. Delinted seed coat fragments produced the lowest recovery. Seed coat fragments carefully removed from ginned lint and added to the processed slivers gave the highest recovery. Averaged SCN recoveries from two AFIS units ranged from 26 to 100% (theoretical). These results helped to explain why the AFIS analysis of SCN counts in processed cotton is lower than by the microscopic analysis.

Characterization of cottonseed protein isolate as a paper additive

ABSTRACT There is current interest in using agro-based biopolymers in industrial applications. Because cottonseed protein is abundantly available, it would be useful to explore its feasibility as a polymeric additive and possible substitute for petroleum-based materials. In this work, we studied cottonseed protein isolate as a paper additive and observed its effects on the paper’s dry and wet strength. The tensile strength of paper was found to vary with the amount of the protein applied. By application of an 11% protein solution to the paper, the dry and wet strength increased by 33 and 16% compared with the paper by itself, respectively. The combined use of cottonseed protein and an acid (acetic, adipic, aspartic, and citric acids) to promote adhesion resulted in even greater dry paper strength but not in greater wet paper strength. Thermogravimetric analysis, infrared spectroscopy, and scanning electron microscopic studies suggested that the protein interacted with acid and that both components interacted with paper fibers to produce increased strength.

Gossypolhemiquinone, a dimeric sesquiterpenoid identified in cotton (Gossypium).

The report that the cotton leaf perforator, Bucculatrix thurberiella, is one of the few insect herbivores to attack Gossypium thurberi prompted an investigation of the terpenoids present in the leaves of this wild species of cotton. Members of Gossypium produce subepidermal pigment glands in their leaves that contain the dimeric sesquiterpenoid gossypol as well as other biosynthetically related terpenoids. In addition to gossypol, a previously unknown dimeric sesquiterpenoid, gossypolhemiquinone (GHQ), was identified in trace amounts in G. thurberi, a member of the D genome. Other members of the D genome of Gossypium were subsequently found to contain this compound, but GHQ was not detected in commercial cotton cultivars. When fed to Helicoverpa zea in an artificial diet, GHQ delayed days-to-pupation, reduced pupal weights, and survival to adulthood to a lesser or equal extent than gossypol in comparison to the control diet. However, GHQ had a synergistic effect on survival and days-to-pupation when combined with gossypol at the highest dosage tested (0.18%; 15.5:84.5 GHQ:gossypol). Because gossypol exhibits anti-cancer activity, GHQ was also evaluated for its anti-cancer activity against the National Cancer Institutes 60-Human Tumor Cell Line Screen. Significant inhibitory activity against most of these cell lines was not observed, but the results may offer some promise against leukemia cancer cell lines.