Dennis T. Ray

2,3-Dihydrowithaferin A-3β-O-sulfate, a new potential prodrug of withaferin A from aeroponically grown Withania somnifera

Preparations of the roots of the medicinal plant Withania somnifera (L.) Dunal commonly called ashwagandha have been used for millennia in the Ayurvedic medical tradition of India as a general tonic to relieve stress and enhance health, especially in the elderly. In modern times, ashwagandha has been shown to possess intriguing antiangiogenic and anticancer activity, largely attributable to the presence of the steroidal lactone withaferin A as the major constituent. When cultured using the aeroponic technique, however, this plant was found to produce a new natural product, 2,3-dihydrowithaferin A-3beta-O-sulfate (1), as the predominant constituent of methanolic extracts prepared from aerial tissues. The characteristic bioactivities exhibited by 1 including inhibition of cancer cell proliferation/survival, disruption of cytoskeletal organization and induction of the cellular heat-shock response paralleled those displayed by withaferin A (2). The delayed onset of action and reduced potency of 1 in cell culture along with previous observations demonstrating the requirement of the 2(3)-double bond in withanolides for bioactivity suggested that 1 might be converted to 2 in cell culture media and this was confirmed by HPLC analysis. The abundant yield of 1 from aeroponically cultivated plants, its good aqueous solubility and spontaneous conversion to 2 under cell culture conditions, suggest that 1 could prove useful as a readily formulated prodrug of withaferin A that merits further evaluation in animal models.

Growth and yield characteristics of Lesquerella fendleri as a function of plant density

Abstract There is little information available regarding the response of Lesquerella fendleri to planting density. The effects of plant density on growth, yield, and yield components of L. fendleri were studied in central Arizona at plant populations of 250 000, 500 000, 750 000, 1 000 000, and un-thinned controls of 1 658 000 plants/ha in 1992–1993 and 1 500 000 plants/ha in 1993–1994. In the 1992–1993 tests, increasing density resulted in a significant decrease in plant width and number of branches, while causing an increase in taproot length in populations up to 750 000 plants/ha. There were no effects on seed size; however, density significantly reduced the number of seeds/silique for the 750 000 and 1 658 000 plants/ha treatments. Seed yield/plant and harvest index/plant were significantly reduced with increasing density, but yield/ha increased with density up to 1 000 000 plants/ha. At 1 658 000 plants/ha, shattering was excessive and resulted in loss of seeds. The greatest seed yields/ha were obtained at 750 000 plants/ha and 1 000 000 plants/ha densities. The oil content did not vary significantly from 500 000 to 1 658 000 plants/ha, but treatments in this range produced seeds with significantly higher oil content than the 250 000 plants/ha treatment. Differences in oil yield/ha reflected differences in seed yield/ha. The 1993–1994 harvest did not show significant differences for seed and oil yields among density treatments. Higher temperatures in 1993–1994 were believed to increase the oil content and seed yield in the higher plant densities. Plant densities of 750 000 and 1 000 000 plants/ha are recommended for lesquerella growing areas comparable to central Arizona. At these densities, maximum seed yields should be obtained even with annual temperature fluctuations. Also, maximum plant heights were obtained at these densities, minimizing seed loses during combine harvest.

Lesquerella fendleri seed oil content and composition: Harvest date and plant population effects

Abstract Yield losses of Lesquerella fendleri L. seeds, caused by shattering and seed oil content reduction by excessive desiccation could be minimized by the appropriate choice of harvest dates. However, no information is available on harvest dates to maximize economic returns at any planting density. This research was undertaken to study the effects of harvest dates and plant populations on oil yield and composition in L. fendleri. Mechanically seeded field plots of L. fendleri were thinned at 103 days after sowing (DAS) to density treatments of 250,000, 500,000, 750,000, 1,000,000, and an nonthinned control of 1,500,000 plants/ha. These plants were grown in 1993–1994 at The University of Arizona, Maricopa Agricultural Center in central Arizona, and were harvested at 203, 229, 243, and 264 DAS. Eight fatty acids, which make up 87–96% of mature seed oil, were investigated. These included palmitic (C16: 0), palmitoleic (C16: 1), stearic (C18: 0), oleic (C18: 1), linoleic (C18: 2), linolenic (C18: 3), lesquerolic (C20: 1-OH), and auricolic (C20: 2-OH) fatty acids. Generally, seed at the earliest harvest of 203 DAS had lower concentrations of palmitoleic, stearic, oleic, lesquerolic, and auricolic acids, and lower total oil content than those harvested at 229, 243, and 264 DAS. Also at 203 DAS, the 750,000 plants/ha treatment produced the highest fatty acid content yield, although both this oil and that from the 1,000,000 plants/ha treatment lacked auricolic acid. During this period, the major contributing fatty acids to oil content were oleic, linoleic, linolenic, and lesquerolic. Harvest at 229 DAS resulted in an increase in oil content for all treatments except the 750,000 plants/ha. Lesquerolic acid was the dominant oil for the nonthinned control. At 243 DAS, higher oil contents were observed in the 250,000, 750,000, and 1,000,000 plants/ha densities, with lesquerolic acid again the dominant fatty acid. At final harvest (264 DAS), lesquerolic acid content was virtually the same among the density treatments. No significant differences for seed yield, fatty acid composition or content were found among plant populations. The seed yield averaged over all population densities was 1344 kg/ha with a fatty acid content of 21.9%, giving an oil yield 267 kg/ha (calculated as total fatty acid content).

Heritability of height, width, resin, rubber, and latex in guayule (Parthenium argentatum)

Abstract Plant breeding in guayule, a rubber and latex producing plant, has not been as effective as predicted or desired. A surprisingly large amount of variability has been reported in this apomictic crop for traits such as plant height, width, resin, rubber, and latex contents. This study was designed to calculate the proportion of the total measured variability due to the environment and the proportion due to genetic influences within and between three-released germplasm lines. Plant heights were measured at 1, 2 and 3-years-of-age; plant width, and latex contents at 2 and 3-years-of-age; and resin and rubber content at 2-years-of-age. Broad-sense-heritabilities were estimated for each trait and year by dividing the genotypic by the total variance. To estimate the genetic component of the measured variance for each trait, the environment effects (variance from clonally propagated plants) were subtracted from the total variance (variance from open-pollinated (OP) seed propagated plants). In general, the variances of the means for the measured traits were lower in the clonally propagated plants compared with the apomictic OP seed propagated plants. The heritability estimates calculated for each trait differed from year to year. For instance, heritability for plant height was estimated in line AZ-2 to be 0.84 at 1 year of age; 0.47 at 2 years of age; and 0.0 at 3 years of age. These values imply that a large portion of the observed variation in this line is attributed to genetic effects in the first 2 years of growth. As the plant grows over several seasons, the environment effects compound, masking the genetic effects, making effective selection choices more difficult. Heritability for latex content for the same line was estimated to be 0.97 for the second year and 0.55 for the third year. Selections for the measured traits in this study appear to be most effective during the first and second years of growth, with effectiveness diminishing during the third year. Most selections previous to this study were performed between 3 and 5 years of growth, thus suggesting one reason for the lack of significant progress in most breeding programs.

Improvement in hydroxy fatty acid seed oil content and other traits from interspecific hybrids of three Lesquerella species: Lesquerella fendleri, L. pallida, and L. lindheimeri

Interspecific hybridization offers potential to improve the hydroxy fatty acid (HFA) content of lesquerella. Lesquerella fendleri is currently being developed for cultivation as a potential new industrial oilseed crop because of its seed productivity. However, it has lower HFA content compared to L. pallida and L. lindheimeri. The objectives of this research were to improve the HFA oil content of L. fenderi through interspecific hybridization and to characterize hybrids and successive generations for seed oil fatty acid profile, fertility, seed set and other morphological traits. In this work, three species were successfully hybridized, self-pollinated, and backcrossed. Ovule culture was used in some cases to produce progeny where interspecific hybrids did not produce viable embryos. The traits measured were petal length, ovules per silique, seeds per silique, and weight of 1000 seed. Patterns of leaf trichomes were used to distinguish between parents and hybrids. Seed per silique indicated that autofertility occurred in L. pallida but not in the other two species. HFA oil content of L. fendleri seed were 50.5% compared to 80 and 84% for L. pallida and L. lindheimeri, respectively. The HFA oil content of the hybrids ranged from 57 to 70% in A2 and A3 generations, and the range of values depended on the parents used in the crosses. These measurements will help predict the value of different interspecific crosses for breeding. Segregation for various yield related traits should allow selection for favorable improvements in the HFA trait and in seed yield.

New Crops Breeding: Lesquerella

Lesquerella species contain a seed oil which is approximately 55% lesquerolic acid, a 20-carbon long fatty acid with a single hydroxyl group and double bond, and has a similar hydroxy fatty acid (HFA) profile as castor oil. Large markets exist for hydroxylated oils as feedstocks for lithium greases, polymers in paints and coatings, base stocks for lubricants, nylon-11, hydraulic fluids, and applications in the personal care industry (Roetheli et al. 1992). The hydroxyl group of these oils makes them prime candidates as additives to diesel fuel to improve lubricity (Naughton 1992). Goodrum and Geller (2005) demonstrated that lesquerella oil has superior performance compared to castor, soybean, and rapeseed methyl esters at concentrations as low as 0.25% in reducing wear and damage to diesel engines, primarily with fuel injection systems. Castor oil also contains high amounts of HFAs, but the main HFA, ricinoleic acid, is two carbons shorter than lesquerolic acid, which imparts slightly different physicochemical properties to the oil. Lesquerella could be established as a reliable, domestic oilseed supply of HFAs for a variety of industrial applications (Roetheli et al. 1992) and at the same time provide an alternative crop for farmers and increase local profits. Lesquerella will not replace current commodity crops but instead will be placed in a rotation with these crops, e.g., a 2-year, 3-crop rotation of lesquerella, grain sorghum, and cotton.

Relationship of morphological variables to rubber production in guayule

SummaryThe objective of this study was to find measurable characters to aid the plant breeder in predicting both rubber percentage and rubber yield of guayule (Parthenium argentatum Gray). A field of 234 guayule breeding lines was harvested by clipping in February 1986. Forty-two of these lines were selected for further evaluation on the basis of superior regrowth, rubber content and yield. These 42 lines were reharvested in February 1987. Nineteen variables were evaluated including percent rubber and resin, rubber and resin yield (g/plant), plant height (cm), width (cm), and volume (m3), fresh and dry weight (kg/plant), percent dry weight, stem number, mean stem diameter (cm), total and mean stem circumference (cm), total and mean stem area (cm2), total stem circumference/total stem area (cm-1), and regeneration at two dates. Dry weight (kg/plant) had the best correlation to rubber yield (r=0.85), and plant volume best predicted dry weight (r=0.83). These variables did not correlate as highly with percent rubber. The best statistical regression model for percent rubber included four variables, fresh and dry weight, percent dry weight, and plant volume. This model accounted for 51% of the variation. In comparison, the best regression of rubber yield had an R2=0.85 for plant height and width, volume, and dry weight.

Growth Responses and Nitrogen Uptake by Saltgrass (Distichlis spicata L.), a Halophytic Plant Species, under Salt Stress, Using the 15N Technique

Various saltgrass clones were studied hydroponically, using Hoagland solution, in a greenhouse to evaluate their DM weights and nitrogen uptake under control and salt stress conditions. Treatments included control (no added salt) and plants grown under NaCl salinity. Twelve clones were grown with 4 replications of each treatment in a RCB design trial. Ammonium sulfate, 5.3%15N was used to enrich the plants by adding 5 mg 15N as 22.931 mg (15NH4)2SO4, per liter of the culture solution per day. Plant shoots were harvested weekly, oven-dried at 65°C, and DM weights were recorded. At the last harvest, plant roots were also harvested, oven-dried at 65°C, and DM weights were determined. Harvested plant materials were analyzed for total-N and 15N contents. The results showed non-significant differences in shoot DM weights and total-N and 15N concentrations and contents in salinized plants compared with the controls. Total-N and 15N concentrations of the roots were higher than that of the shoots under either control or saline condition. Overall, due to the high degree of salt tolerance of saltgrass, the results showed generally no difference in nitrogen uptake by most of the clones under salt stress compared with the control plants.

Variation of agronomic characters among and between guayule lines

SummaryA selected group of 42 guayule (Parthenium argentatum Gray) lines were evaluated on the basis of various morphological characteristics and compared to a total population of 234 lines. The objective of this study was to determine the amount of variation that existed both within and among guayule lines over a 2-year period. The parental lines all reproduced apomictically, and as expected, many lines exhibited a high degree of uniformity for every measured character. However, many other lines were extremely variable for varying numbers of characters. This appears to be due to varying degrees and combinations of apomeiosis, thus establishing a long-term source of genetic variation. The high amount of variability existing among and within these guayule lines indicates that significant progress by selection for both rubber percentage and yield is feasible.

Parent-offspring relationships in apomictic guayule

The genetic base of guayule (Parthenium argentatum Gray) germ plasm that is available is narrow; yet, significant variability has been described within this germ plasm. This variability is surprising because guayule reproduces by facultative apomixis (asexual reproduction by seed), but evidence suggests that progress through selection is feasible. It has been hypothesized that this variation has arisen through periodic sexual reproduction; however, this has not been quantified. This study was designed to describe and compare eight components of yield and the expression of two isozyme systems in twenty parent plants and half-sib, open-pollinated families from each parent. Extensive variation for all characters was found among both the parents and the progeny, with fresh and dry weights being highly correlated to rubber yield. Parent-offspring regressions were not significantly different from zero. This implies low heritabilities for the major components of yield in guayule, and we believe that family selection, rather than single-plant selection, may generate higher-yielding and more genetically uniform lines at an escalated rate than single-plant selection. Periodic sexuality resulting in genetic variation was estimated by differences in isozyme expression within the half-sib families. Fifteen of the twenty progeny families were polymorphic for either esterase or peroxidase or both isozymes. This amount of sexuality is significant, and guayule must be considered an open-pollinated crop. We conclude from this study that we should rethink our breeding procedures to include controlled pollination and family selection.