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Cet article s’intéresse aux actes de condamnation de l’être (Laforest & Moïse 2013) dans le contexte de l’interaction parent-enfant, adressés par le parent maltraitant à son enfant et rapportés par cette dernière ou ce dernier dans des témoignages anonymes publiés sur le compte Instagram francophone Parents toxiques. L’étude porte sur un échantillon de dix témoignages publiés entre mai 2020 et juillet 2021. L’objectif est, d’une part, de caractériser les modalités du traitement de l’altérité qui contribuent à une perception de l’interaction comme inappropriée ou maltraitante; d’autre part, d’interroger si le rapport d’autorité constitutif de l’interaction parent-enfant est perturbé dans et par les actes de condamnation de l’être, et si oui, comment. À cette fin, l’étude s’organise en trois temps: le premier volet explicite la méthodologie d’identification et d’analyse des actes de condamnation rapportés; le deuxième met en évidence les ressorts discursifs des actes de condamnation de l’être rapportés par les autrices et l’auteur des témoignages; le dernier volet s’attache à caractériser les motifs qui sous-tendent la construction de l’être comme fautif, et les implications de ceux-ci en termes de rapport de places. Les analyses menées montrent que la condamnation de l’être puise sa force pragmatique dans sa légitimation, construite en discours et interaction, par l’appui sur des normes présentées comme universelles et la mise en saillance du préjudice causé par l’autre. Construire l’être comme fautif consiste, à différents degrés, à essentialiser et rétrograder l’autre ainsi qu’à faire un amalgame entre sa valeur intrinsèque et sa valeur pour soi.

Tralokinumab is the first selective interleukin 13 inhibitor approved for moderate to severe atopic dermatitis. This article reports the findings of a comprehensive literature review and extensive economic analysis to assess tralokinumab's safety, effectiveness, and cost. Evidence synthesis involved evaluating comparative effectiveness and conducting economic sensitivity analyses. This review was prepared by the University of Connecticut School of Pharmacy Academy of Managed Care Pharmacy (AMCP) Student Chapter. The student author group won the AMCP National Pharmacy and Therapeutics competition for their tralokinumab product review in March 2023.

Brown rice is nutritionally superior to white rice, yet oil rancidity can be problematic during processing and storage regarding sensory attributes. Germinating brown rice is known to generally increase some health-promoting compounds. In response to increasing the consumption of plant-based beverages, we sprouted unstabilized brown rice, using green technologies and saccharification enzymes for value-added beverages. 'Rondo' paddy rice was dehulled, sorted and germinated, and beverages were produced and compared against non-germinated brown and white brewers rice beverages. The preliminary germinated brown rice beverage contained significantly higher concentrations of total lipids, diacylglycerols, triacylglycerols, free sterols, phytosterol esters and oryzanols than both non-germinated brown and white rice beverages. White rice beverages had significantly higher free fatty acids. Significant lipid losses occurred during sieving, yet novel germinated brown rice beverages contained appreciable levels of valuable health-beneficial lipids, which appeared to form natural emulsions. Further pilot plant investigations should be scaled-up for pasteurization and adjusted through emulsification to ameliorate sieving losses.

Current evidence indicates that foods with added plant sterols or stanols can lower serum levels of low-density lipoprotein cholesterol. This review summarizes the recent findings and deliberations of 31 experts in the field who participated in a scientific meeting in Winnipeg, Canada, on the health effects of plant sterols and stanols. Participants discussed issues including, but not limited to, the health benefits of plant sterols and stanols beyond cholesterol lowering, the role of plant sterols and stanols as adjuncts to diet and drugs, and the challenges involved in measuring plant sterols and stanols in biological samples. Variations in interindividual responses to plant sterols and stanols, as well as the personalization of lipid-lowering therapies, were addressed. Finally, the clinical aspects and treatment of sitosterolemia were reviewed. Although plant sterols and stanols continue to offer an efficacious and convenient dietary approach to cholesterol management, long-term clinical trials investigating the endpoints of cardiovascular disease are still lacking.

Phytosterols (plant sterols) occur in the cells of all plants. They are important structural components that stabilize the biological membranes of plants. Sterols can occur in the "free" unbound form or they can be covalently bound via an ester or glycosidic bond. Since our previous 2002 review on phytosterols and phytosterol conjugates, phytosterol glucosides have been found to be important structural components in the lipid rafts of the plasma membrane of plant cells, where they are thought to be essential to the function of plasma membrane enzymes and perhaps other proteins. Phytosterols also serve as precursors in the synthesis of important bioactive compounds such as steroidal saponins, steroidal glycoalkaloids, phytoecdysteroids, and brassinosteroids. Methods for the analysis of phytosterols range from traditional gas chromatography of free phytosterols to modern sophisticated forms of mass spectrometry which have been used for the new field of sterol lipidomics, sometimes called "sterolomics." Phytosterol-enriched functional foods first appeared about twenty years ago and many clinical studies have confirmed the low density lipoprotein (LDL) cholesterol-lowering properties of various types of phytosterols. In recent years additional clinical studies and more than ten important meta-analyses have provided insights to better understand the cholesterol-lowering and other biological effects of plant sterols., (Published by Elsevier Ltd.)

The aim of this study was to determine if the compositional difference between grain sorghum and corn impact ethanol yields and coproduct value when grain sorghum is incorporated into existing corn ethanol facilities. Fermentation properties of corn and grain sorghum were compared utilizing two fermentation systems (conventional thermal starch liquefaction and native starch hydrolysis). Fermentation results indicated that protease addition influenced the fermentation rate and yield for grain sorghum, improving yields by 1-2% over non-protease treated fermentations. Distillers Dried Grains with Solubles produced from sorghum had a statistically significant higher yields and significantly higher protein content relative to corn. Lipid analysis of the Distillers Dried Grains with Solubles showed statistically significant differences between corn and sorghum in triacylglycerol, diacylglycerol and free fatty acid levels., (Published by Elsevier Ltd.)

All fruits, vegetables, grains and other plant materials contain small amounts of plant sterols, which are essential for the function of the biological membranes in living cells. The average human consumption of plant sterols has been estimated to be about 150-350 mg/day and trace amounts of stanols (which are defined as saturated sterols such as sitostanol), but this number varies regionally and is higher for vegetarians. When consumed in the diet, plant sterols reduce the levels of serum cholesterol. In 1995 the first functional food product, Benecol spread (enriched in plant stanol fatty acid esters), was developed by Raisio and marketed, first in Finland and then globally. Since then many other functional food products have been developed and are now available globally. In addition to stanol esters, other functional food products contain plant sterol esters and/or free (unesterified) plant sterols and stanols. In essentially all of the current functional foods that are enriched in sterols and stanols, the feedstock from which the sterols and stanols are obtained is either tall oil (a byproduct/coproduct of the pulping of pine wood) or vegetable oil deodorizer distillate (a byproduct/coproduct of the refining of vegetable oils).

Candida bombicola is well-studied for the production of a biosurfactant, the sophorolipids. In this paper, the cloning of a glucosyltransferase gene using polymerase-chain-reaction (PCR) technique is described. Degenerative primer-pairs were first designed based on the highly conserved amino-acid sequences of several selected yeast glucosyltransferases. Using these primers, an amplified sequence (amplicon) of 700 base-pair from C. bombicola was obtained and subsequently sequenced. Based on the sequence of this amplicon, additional target-specific PCR primers were designed for use in subsequent rounds of 3'- and 5'-extension using DNA walking technique to eventually obtain a C. bombicola genomic sequence containing an open-reading-frame putatively identified as a glucosyltransferase (gtf-1). The gene was subcloned in Saccharomyces cerevisiae for expression and functional characterization. Quantitative RT-PCR confirmed the expression of gtf-1 in the recombinant S. cerevisiae. In vitro assay with the sonicated cells of the recombinant yeast confirms the presence of glucosylation activity on sterol and hydroxy fatty acid substrates. This study reports for the first time the cloning and characterization of a broad-specificity lipid glucosylation gene from C. bombicola, and the functional activity of its gene product., (Published by Elsevier B.V.)

Recent breeding advances have led to the development of several barley lines and cultivars with significant reductions (50% or greater) in phytate levels. Low-phytate (LP) grain is distinguished by containing not only a reduced level of phytate P but also an increased level of inorganic P, resulting in greater bioavailability of P and mineral cations in animal diets. It is important to determine whether other nutritional characteristics are altered by breeding for the low-phytate trait. This study was designed to investigate if breeding for reduced phytate content in barleys had any effect on the contents of other attributes measured by comparing mean and range values of the levels of protein, oil, ash, total carbohydrate, starch, and β-glucan, fatty acid composition, and levels of tocopherols and tocotrienols between five LP and five normal-phytate barleys grown in three Idaho locations. Results show that only the phytate level in the LP group was substantially lower than that of the normal-phytate group and that all other attributes measured or calculated were substantially equivalent between the two groups of barleys. Therefore, the phytate level had little effect on the levels of protein, oil, ash, total carbohydrate, starch, and β-glucan, fatty acid composition, and levels of tocopherols and tocotrienols in barley seeds.

This research focuses on the overall extraction process of alkylresorcinols (ARs) from uncooked grains and baked products that have been processed with wheat, corn, rice, and white flour. Previously established extraction methods developed by Ross and colleagues, as well as a semiautomated method involving accelerated solvent extraction (ASE), were applied to extract ARs within freshly ground samples. For extraction of alkylresorcinols, nonpolar solvents such as ethyl acetate have been recommended for the extraction of uncooked foods, and polar solvents such as 1-propanol:water (3:1 v/v) have been recommended for the extraction of baked foods that contain rye, wheat, or other starch-rich grains. A comparison of AR extraction methods has been investigated with the application of gas chromatography and a flame ionization detector (GC-FID) to quantify the AR content. The goal of this research was to compare the rapid accelerated solvent extraction of the alkylresorcinols (ASE-AR) method to the previous manual AR extraction methods. Results for this study as well as the investigation of the overall efficiency of ASE-AR extraction with the use of a spiking study indicated that it can be comparable to current extraction methods but with less time required. Furthermore, the extraction time for ASE (approximately 40 min) is much more convenient and less tedious and time-consuming than previously established methods, which range from 5 h for processed foods to 24 h for raw grains.

In this study, the effect of the 80% ethanolic extract of corn bran (EECB) on inhibition of nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) expression in lipopolysaccharide (LPS)-stimulated Raw 264.7 cells was investigated. The EECB inhibited LPS-induced NO production and iNOS expression in a dose-dependent manner. Four hydroxycinnamic acid derivatives (HADs), including two free cinnamic acids, p-coumaric acid (CA) and ferulic acid (FA), and their conjugate phenolic amides, p-dicoumaroyl-putrescine (DCP) and diferuloylputrescine (DFP), were found to be present in the EECB by LC-MS analysis, and DFP (378.66 μg/g) was the predominant phenolic compound, followed by DCP (7.83 μg/g)>CA (5.58 μg/g)>FA (1.84 μg/g). The four HADs significantly inhibited NO production and iNOS expression in a dose-dependent manner. Among the four HADs tested, DFP showed the most potent inhibition on NO production and iNOS mRNA and protein expression, followed by DCP>FA ≥ CA. DFP also exhibited the strongest inhibition on LPS-induced iNOS and NF-κB luciferase activity, which was followed by DCP ≥ FA (CA)>CA (FA). Thus, these results suggest that phenolic amides in the corn bran may be a potential source of natural anti-inflammatory agents., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Phytosteryl glycosides occur in natural foods but little is known about their metabolism and bioactivity. Purified acylated steryl glycosides (ASG) were compared with phytosteryl esters (PSE) in mice. Animals on a phytosterol-free diet received ASG or PSE by gavage in purified soybean oil along with tracers cholesterol-d(7) and sitostanol-d(4). In a three-day fecal recovery study, ASG reduced cholesterol absorption efficiency by 45 ± 6% compared with 40 ± 6% observed with PSE. Four hours after gavage, plasma and liver cholesterol-d(7) levels were reduced 86% or more when ASG was present. Liver total phytosterols were unchanged after ASG administration but were significantly increased after PSE. After ASG treatment both ASG and deacylated steryl glycosides (SG) were found in the gut mucosa and lumen. ASG was quantitatively recovered from stool samples as SG. These results demonstrate that ASG reduces cholesterol absorption in mice as efficiently as PSE while having little systemic absorption itself. Cleavage of the glycosidic linkage is not required for biological activity of ASG. Phytosteryl glycosides should be included in measurements of bioactive phytosterols.

Evidence going as far back as the early part of the 20th century suggests that both light and chloroplast function may play key roles in host susceptibility to viruses. Despite the long history of such work, confirmation of these phenomena and a determination of the underlying mechanisms remain elusive. Here, we revisited these questions using modern imaging technologies to study the susceptibility of Nicotiana benthamiana to Turnip mosaic virus (TuMV). We found that both light deficiency and photosystem impairment increased the susceptibility of N. benthamiana to TuMV infection. Time-lapse photography studies indicated that, under these conditions, rub-inoculated plants exhibited greater numbers of infection foci and more rapid foci development. The rate of systemic movement was also accelerated though cell-to-cell movement appeared unchanged. Inhibition of salicylic acid (SA)-mediated defense responses is not likely responsible for changes in susceptibility because SA and pathogen response-1 gene induction were not affected by light deficiency or chloroplast impairment and treatment of plants with SA had no measureable impact on TuMV infection. Taken together, these data suggest that both light and optimal chloroplast function influence virus infection either by limiting the cellular resources needed by TuMV to establish replication complexes or the host's ability to activate SA-independent defenses.

Hulless barley kernels were sequentially abraded to achieve 4%, 8%, 16%, 24%, 32%, and 40% removal. Abraded fines, kernels, and ground kernels were stored at 35 degrees C and 75% relative humidity for 3 wk. Stored samples were extracted and the levels of oil, free phytosterols, tocopherols (Ts), and tocotrienols (T3s) were analyzed and compared with freshly abraded fractions. The results revealed that oil, sterols, and Ts were concentrated in the outer layers, particularly in the germ layer. In whole kernels, homologues of both Ts and T3s showed the same ranking order in concentrations as alpha > gamma > beta > delta. The homologue composition of Ts remained the same but that of T3s changed across the kernel. The %T3 in total tocols increased in fractions with increasing endosperm tissue. Storage caused no change in oil and Ts but significant changes in sterols and T3s. The changes were differential among T3 isomers, with alpha-T3 decreasing and delta-T3 increasing. The degradation of alpha-T3 was accelerated in fractions with more endosperm tissue. Grinding kernel samples before storage accelerated sterol degradation but had a limited effect on changes of T3s. A 2nd experiment using a different hulless barley line and ambient storage for 6 mo confirmed all the findings except that the changing trend for sterols was inconsistent. These results provide practical information to those who wish to produce a barley fraction enriched with a particular functional lipid and maintain stability of their products.

Oat kernels were extracted with methanol, and glycolipid-enriched fractions were prepared using silica solid phase extraction. Using direct infusion electrospray ionization (ESI) tandem mass spectrometry (MS), high performance liquid chromatography (HPLC)-ESI-MS, and HPLC-atmospheric pressure chemical ionization (APCI)-MS, we confirmed previous reports that digalactosyldiacylglycerol (DGDG) was the most abundant glycolipid in oat kernels and confirmed a previous report of the presence of a DGDG mono-estolide in oat kernels. In the current study we also identified several additional natural galactolipid estolides: two new DGDG estolides (di- and tri-estolides), two trigalactosyldiacylglycerol (TriGDG) estolides (mono- and di-estolides), and one tetragalactosyldiacylglycerol (TetraGDG) estolide (mono-estolide). The levels of total galactolipid estolides in oat kernels were estimated to be about 29% of the total glycolipid fraction. To our knowledge, this report is the first evidence of natural di- and tri-estolides of polar lipids.

Hominy feed is a low value ($83.7/metric ton) coproduct of the corn dry milling process that accounts for nearly 35% of the starting corn quantity. The average composition of hominy feed on a dry basis is 56.9% starch, 25.2% neutral detergent fiber, 11.1% protein, and 5.3% fat. Starch in hominy feed can be fermented to ethanol thus increasing its levels of protein and fat. The increase in protein and fat percentages may increase the market competitiveness and price of hominy feed. Hydrolysis and fermentation were performed on nine hominy feed samples collected from three corn dry milling plants in the USA. The original hominy feed samples and postfermentation solids were analyzed for starch, protein, fat, and fiber content. Compared to the original hominy feed, the percentage increase in protein, fat and fiber in postfermentation solids of nine samples ranged from 10.4 to 21.3, 6.78 to 10.6, and 12.6 to 28.7% (dry basis), respectively. Ethanol yields varied from 271.7 to 380.2 l/metric ton for the nine hominy feed samples. These results indicate that the value of hominy feed as an animal feedstock can potentially be increased with fermentation and can produce more profit per metric ton than currently being derived by its sale as a low protein feed ingredient.

Bioprocesses were developed to enhance the value of proteins from deoiled corn germ. Proteins were hydrolyzed with trypsin, thermolysin, GC 106, or Flavourzyme to generate the bioactive peptide sequences. At an enzyme to substrate ratio of 1:100, protein hydrolysis of wet-milled germ was greatest using thermolysin followed by trypsin, GC 106, and Flavourzyme. For the dry-milled corn germ, protein hydrolysis was greatest for GC 106 and least for Flavourzyme. Electrophoretic patterns indicated that the hydrolysis conditions used were adequate for generating low molecular weight peptides for both germs. Unhydrolyzed dry- and wet-milled corn germ did not appear to contain angiotensin I converting enzyme (ACE)-inhibitory peptides. After hydrolysis with trypsin, thermolysin, and GC 106 but not Flavourzyme, ACE inhibition was observed. ACE inhibition was greatest for the GC 106 hydrolysate for both wet- and dry-milled corn germ. Denaturing the protein with urea before hydrolysis, in general, increased the amount of ACE-inhibitory peptides found in the hydrolysate. Membrane fractionations of both the wet- and dry-milled hydrolysates indicated that most of the ACE-inhibitory peptides were in the <1 kDa fraction. Examination of the control total protein extracts (before treatment with proteases) from wet- and dry-milled germ revealed that neither had ACE-inhibitory properties. However, when both total corn germ control protein extracts were fractionated, the <1 kDa fraction of wet-milled corn germ proteins exhibited ACE inhibition, whereas the comparable low molecular weight fraction from dry-milled corn germ did not.

Objective: The aim of this study was to investigate the cholesterol-lowering mechanisms of corn fiber oil (CFO), ferulate phytostanyl esters (FPEs) and parent compounds of FPE, including sitostanol and ferulic acid, in hamsters., Method: Seventy male Golden Syrian hamsters were randomly assigned to six experimental diets for 4 weeks: (1) cornstarch-casein-sucrose-based control diet (control); and (2) control diet plus 0.1% (wt/wt) cholesterol (cholesterol-control). The remaining four groups were given cholesterol-control diet with: (3) 10% (wt/wt) CFO; (4) 0.5% (wt/wt) sitostanol; (5) 0.23% (wt/wt) ferulic acid; and (6) 0.73% (wt/wt) FPE. At the end of dietary intervention, total plasma cholesterol, high-density lipoprotein cholesterol and triglyceride concentrations were determined. Parameters of cholesterol kinetics, including cholesterol absorption and synthesis, as well as mRNA expression of sterol transporters such as Niemann-Pick C1 like 1 (NPC1L1), ATP-binding cassette G5 (ABCG5) and ABCG8, were assessed., Results: Supplementation with CFO decreased (P<.0001) plasma total cholesterol levels by 29% as compared with the cholesterol-control group, while FPE and sitostanol reduced (P<.02) cholesterolemia by 15% and 14%, respectively. CFO and sitostanol decreased (P<.05) cholesterol absorption by 24% compared to the cholesterol-control group. Dietary intervention did not alter the intestinal gene expression of ABCG5, ABCG8 and NPC1L1., Conclusion: The present results show that the CFO-induced and sitostanol-induced decrease in cholesterol absorption is independent of intestinal enterocyte sterol transporters such as ABCG5, ABCG8 and NPC1L1 in hamsters.

Brown rice is a valuable source of lipid-soluble antioxidants including ferulated phytosterols (i.e., gamma-oryzanol), tocopherols, and tocotrienols. To evaluate the impact of temperature on the accumulation of these compounds, seeds from six different rice lines grown to maturity in replicate greenhouses in Gainesville, FL, were analyzed. The lines represented Oryza sativa indica, O. sativa japonica, and Oryza glaberrima of different origins. Temperatures were maintained near ambient at one end of each greenhouse and at approximately 4.5 degrees C above ambient at the other end. gamma-Oryzanols, tocopherols, and tocotrienols were extracted from whole seed (i.e., brown rice) and analyzed by HPLC. Tocotrienols and tocopherols varied widely between lines but changed only slightly with respect to temperature. In general, the proportions of alpha-tocotrienol and/or alpha-tocopherol increased at elevated temperature, whereas gamma-tocopherol and gamma-tocotrienol decreased. Six gamma-oryzanol peaks, identified on the basis of absorbance maxima at 330 nm and HPLC-mass spectrometry, were quantified. The most abundant component was 24-methylenecycloartanyl ferulate, present at 40-62% of total. Its levels increased 35-57% at elevated temperature in five of six lines, accounting for most of the change in total gamma-oryzanol. The results suggest that the physiological action of individual ferulated phytosterols should be investigated because their relative proportions in gamma-oryzanol can change.

The antioxidant activity of three major polyamine conjugates, N,N'-dicoumaroyl-putrescine (DCP), N-p-coumaroyl-N'-feruloylputrescine (CFP), and N,N'-diferuloyl-putrescine (DFP) isolated from corn bran, and their related hydroxycinnamic acids, p-coumaric acid and ferulic acid, were evaluated by three antioxidant in vitro assay systems, including 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical and superoxide and hydroxyl radicals generated by enzymatic and nonenzymatic reactions. Additionally, five phenolic compounds were evaluated for melanogenesis inhibitory activity using mushroom tyrosinase and B16 melanoma cells. Most of the phenolic compounds significantly scavenged DPPH, superoxide, and hydroxyl radicals in a dose-dependent manner. Particularly, DFP showed potent DPPH (IC50 = 38.46 microM) and superoxide (IC50 = 291.62 microM) radical scavenging activities, while DCP exhibited the strongest hydroxyl radical scavenging activity (IC50 = 120.55 microM). CFP also exerted moderate DPPH, superoxide, and hydroxyl radical scavenging activities. Meanwhile, DCP (IC50 = 181.73 microM) showed potent tyrosinase inhibitory activity toward l-tyrosine as the substrate, whereas DFP (IC50 = 733.64 microM) significantly inhibited melanin synthesis in B16 melanoma cells. These current results indicate that these three polyamine conjugates from corn bran may be useful potential sources of natural antioxidants and skin-whitening agents.

Corn fiber gum (CFG) is a hemicellulose (arabinoxylan)-enriched fraction obtained by the extraction of corn bran/fiber using a proprietary alkaline hydrogen peroxide process. When purified CFG prepared by this process was hydrolyzed with more concentrated base (1.5 N methanolic KOH at 70 degrees C for 1 hour), considerable amounts of hydroxycinnamic acids (up to 0.015% of mainly ferulic acid) and lipids (up to 0.43%) were released. The released phenolic acids and lipids were identified and quantified using high-performance liquid chromatography (HPLC) with detection by both UV and evaporative light-scattering detection (ELSD). During the wet milling of corn, two types of corn fiber are produced: coarse fiber, which is primarily from pericarp, and fine fiber, which is from the endosperm. The total phenolic acid content in CFGs purified from coarse corn fiber (pericarp fiber) is comparatively higher than that purified from fine corn fiber (endosperm fiber). It was also determined that the purified CFG samples contained significant amounts of strongly associated proteins, from 2 to 5% by weight. The presence of these phenolic acids, lipids, and proteins strongly associated or bound to CFG may contribute to its excellent ability to emulsify oil-in-water emulsions.

We previously reported that heat pretreatment of corn fiber (150 degrees C, 1 h) caused a tenfold increase in the levels of extractable gamma-tocopherol. The current study was a reinvestigation of the previous effect, using improved methods (HPLC with fluorescence detection, diode-array UV detection, and mass spectrometry) for tocol analysis. Heat pretreatment did not cause an increase in the levels of any of the tocopherols or tocotrienols in corn fiber oil, but lowered the levels of three of the tocols and had no effect on the levels of the other two tocols. Heat pretreatment of corn germ had a similar effect. UV and mass spectra indicated that the peak that we had identified as gamma-tocopherol in our previous report was probably a mixture of oxidation products of triacylglycerols. Thus, heat treatment of corn germ or other corn-oil containing fractions at high temperatures leads to decreases in gamma-tocopherol, gamma-tocotrienol, and delta-tocotrienol and to the production of triacylglycerol oxidation products.

Because most lipid extracts are a mixture of saturated and unsaturated molecules, the most successful strategies for the quantitative analysis of lipids have involved the use of so-called "mass" or universal detectors such as flame ionization detectors and evaporative light scattering detectors. Recently a new type of HPLC "mass" detector, a charged aerosol detector (CAD), was developed and is now commercially available. This detection method involves nebulizing the HPLC column effluent, evaporating the solvents, charging the aerosol particles, and measuring the current from the charged aerosol flux. In the present study, the CAD was evaluated with several normal phase and reverse phase HPLC methods commonly used for the quantitative analysis of lipid classes and lipid molecular species. The CAD detected common lipids such as triacylglycerols, diacylglycerols, glycolipids, phospholipids, and sterols. Lower molecular weight lipids such as free FA had smaller peak areas (50-80% lower). FAME were not detected by the CAD, probably because they were completely evaporated and did not form aerosol particles. The minimum limits of detection of the CAD with lipids varied with different mobile phase solvents. Using solvent systems that were predominantly hexane, the minimum limits of detection of triacylglycerols, cholesterol esters, and free sterols were about 1 ng per injection and the mass-to-peak area ratio was nearly linear from the range of about 1 ng to about 20 mg per injection. Three other solvents commonly used for HPLC lipid analysis (methanol, isopropanol, and acetonitrile) caused higher levels of background noise and higher minimum limits of detection. These experiments indicate that the CAD has the potential to become a valuable tool for the quantitative HPLC analysis of lipids. Long-term studies are needed to evaluate full instrument performance.

To identify high-valued coproducts from commercially processed corn germ, it was necessary to determine the effect of processing conditions on corn germ proteins. We found that significantly less protein was extracted from commercial wet-milled as compared to dry-milled corn germ using Tris, sodium dodecyl sulfate (SDS) buffer containing 14 mM 2-mercaptoethanol at 100 degrees C for 10 min. SDS-polyacrylamide gel electrophoresis (PAGE) revealed a number of proteins with molecular masses ranging from approximately 10 to 66 kDa for the dry-milled corn germ as compared to only a few significant protein bands centered around 23 kDa in the wet-milled corn germ. The protein content of the wet- and dry-milled corn germ was approximately the same; however, nonprotein nitrogen values were significantly greater for the wet-milled than for the dry-milled germ. The distribution of fractionated germ protein freshly excised from the embryo of yellow dent corn kernels was more similar to that of dry-milled than wet-milled corn. SDS-PAGE of laboratory preparations of wet-milled corn germ more closely resembled commercial dry- than wet-milled corn germ, which could be attributed to limited microbial growth during steeping in the laboratory preparations.

The lipid profiles of cotton fiber cells were determined from total lipid extracts of elongating and maturing cotton fiber cells to see whether the membrane lipid composition changed during the phases of rapid cell elongation or secondary cell wall thickening. Total FA content was highest or increased during elongation and was lower or decreased thereafter, likely reflecting the assembly of the expanding cell membranes during elongation and the shift to membrane maintenance (and increase in secondary cell wall content) in maturing fibers. Analysis of lipid extracts by electrospray ionization and tandem MS (ESI-MS/MS) revealed that in elongating fiber cells (7-10 d post-anthesis), the polar lipids-PC, PE, PI, PA, phosphatidylglycerol, monogalactosyldiacylglycerol, digalactosyldiacylglycerol, and phosphatidylglycerol-were most abundant. These same glycerolipids were found in similar proportions in maturing fiber cells (21 dpa). Detailed molecular species profiles were determined by ESI-MS/MS for all glycerolipid classes, and ESI-MS/MS results were consistent with lipid profiles determined by HPLC and ELSD. The predominant molecular species of PC, PE, PI, and PA was 34:3 (16:0, 18:3), but 36:6 (18:3,18:3) also was prevalent. Total FA analysis of cotton lipids confirmed that indeed linolenic (18:3) and palmitic (16:0) acids were the most abundant FA in these cell types. Bioinformatics data were mined from cotton fiber expressed sequence tag databases in an attempt to reconcile expression of lipid metabolic enzymes with lipid metabolite data. Together, these data form a foundation for future studies of the functional contribution of lipid metabolism to the development of this unusual and economically important cell type.

Chemical analyses of ancient organics absorbed into pottery jars from the early Neolithic village of Jiahu in Henan province in China have revealed that a mixed fermented beverage of rice, honey, and fruit (hawthorn fruit and/or grape) was being produced as early as the seventh millennium before Christ (B.C.). This prehistoric drink paved the way for unique cereal beverages of the proto-historic second millennium B.C., remarkably preserved as liquids inside sealed bronze vessels of the Shang and Western Zhou Dynasties. These findings provide direct evidence for fermented beverages in ancient Chinese culture, which were of considerable social, religious, and medical significance, and help elucidate their earliest descriptions in the Shang Dynasty oracle inscriptions.

A mixture of diferuloylputrescine/p-coumaroylferuloylputrescine (85:15, w/w) demonstrated inhibitory activity against aflatoxin B1 biosynthesis in Aspergillus flavus isolate AF13. Inhibition was concentration dependent, with a 50% effective dose of 30 microg of diferuloylputrescine/p-coumaroylferuloylputrescine per milliliter of medium. Aflatoxin inhibition levels of up to 93% were achieved using this conjugated polyamine material. This diconjugated polyamine mixture did not display inhibitory effects on A. flavus growth (mycelial weight) at any of the concentrations tested. A survey of hand-dissected corn (Zea mays) kernel tissues, including endosperm, germ, pericarp, and wax, revealed that the highest concentrations of these conjugated polyamines were localized in the pericarp of the seed. Analysis of a number of corn accessions did not reveal a correlation between diferuloylputrescine/ p-coumaroylferuloylputrescine concentration and resistance/susceptibility to A. flavus infection. The localization of these diconjugated polyamine components in the pericarp, which functions as a physical barrier and surrounds the internal food storage reserves, suggests a defensive role for these materials., (Copyright 2004 American Chemical Society)

Because of the positive health effects of phytosterols, phytosterol-enriched foods and foods containing elevated levels of natural phytosterols are being developed. Phytosterol contents in cereals are moderate, whereas their levels in the outer layers of the kernels are higher. The phytosterols in cereals are currently underutilized; thus, there is a need to create or identify processing fractions that are enriched in phytosterols. In this study, pearling of hulless barley and rye was investigated as a potential process to make fractions with higher levels of phytosterols. The grains were pearled with a laboratory-scale pearler to produce pearling fines and pearled grains. Lipids were extracted by accelerated solvent extraction, and nonpolar lipids were analyzed by normal-phase HPLC with ELSD and UV detection. Total sterol analyses were performed by GC. After a 90-s pearling, the amounts of pearling fines from hulless barley and rye were 14.6 and 20.1%, respectively, of the original kernel weights. During pearling, higher levels of phytosterols and other lipids were fractionated into the fines. The contents of free sterols and sterols esterified with FA in the fines were at least double those in the whole grains. Pearling fines of hulless barley and rye contained >2 mg/g phytosterol compounds, which makes them a good source of phytosterols and thus valuable raw materials for health-promoting foods.

All fruits, vegetables, and grains contain phytosterols. Numerous clinical studies have documented that phytosterols lower LDL-cholesterol levels and thereby reduce the risk of cardiovascular disease. Most experts believe that the cholesterol-lowering mechanism of phytosterols requires that they be in their "free" form. In addition to their occurrence in the free form, phytosterols also occur as four common phytosterol conjugates: (i) fatty acyl esters, (ii) hydroxycinnamate esters, (iii) steryl glycosides, and (iv) fatty acylated steryl glycosides. This study was undertaken to investigate the extent of hydrolysis of four common phytosterol conjugates by mammalian digestive enzymes (cholesterol esterase and pancreatin, a mixture of pancreatic enzymes) and for comparison purposes, by KOH. Two types of purified hydroxycinnamate esters (sitostanyl ferulate and oryzanol, a mixture of hydroxycinnamate esters purified from rice bran oil) were hydrolyzed by cholesterol esterase and by pancreatin. Both cholesterol esterase and pancreatin hydrolyzed the phytosteryl esters in two functional food matrices, and they hydrolyzed the hydroxycinnamate esters in corn fiber oil. This is the first report to demonstrate that phytostanyl ferulate esters (which are present at levels of 3-6% in corn fiber oil) are hydrolyzed by pancreatic cholesterol esterase. It is also the first report that pancreatin contains enzymes that hydrolyze the fatty acyl moiety of fatty acylated steryl glycoside, converting it to steryl glycoside. Pancreatin had no effect on steryl glycosides. The ability of pancreatin to hydrolyze three other types of lipid conjugates was also evaluated. Phospholipids were completely hydrolyzed. About half of the galactolipids were hydrolyzed, and less than 10% of the polyamine conjugates were hydrolyzed. The extents of hydrolysis of phytosteryl esters by base (saponification) were also studied, and conditions commonly used for the saponification of acyl lipids (1.5 N methanolic KOH, 30 min at 70 degrees C), were found to result in a nearly 100% hydrolysis of TAG but only about 35-45% hydrolysis of the phytosteryl fatty acyl esters or phytosteryl hydroxycinnamate esters.

Approximately 9% of the 9.7 billion bushels of corn harvested in the United States was used for fuel ethanol production in 2002, half of which was prepared for fermentation by dry grinding. The University of Illinois has developed a modified dry grind process that allows recovery of the fiber fractions prior to fermentation. We report here on conversion of this fiber (Quick Fiber [QF]) to ethanol. QF was analyzed and found to contain 32%wt glucans and 65%wt total carbohydrates. QF was pretreated with dilute acid and converted into ethanol using either ethanologenic Escherichia coli strain FBR5 or Saccharomyces cerevisiae. For the bacterial fermentation the liquid fraction was fermented, and for the yeast fermentation both liquid and solids were fermented. For the bacterial fermentation, the final ethanol concentration was 30 g/L, a yield of 0.44 g ethanol/g of sugar(s) initially present in the hydrolysate, which is 85% of the theoretical yield. The ethanol yield with yeast was 0.096 gal/bu of processed corn assuming a QF yield of 3.04 lb/bu. The residuals from the fermentations were also evaluated as a source of corn fiber oil, which has value as a nutraceutical. Corn fiber oil yields were 8.28%wt for solids recovered following pretreatment.

Plant sterols (phytosterols) have been shown to possess serum cholesterol-lowering properties. In recent years, several phytosterol-enriched functional food products have been developed and marketed. Some phytosterol products contain common unsaturated sterols and some contain a subset of phytosterols called phytostanols (saturated sterols, also called plant stanols). Current methods for the quantitative analysis of plant sterols are labor intensive and require sophisticated gas or liquid chromatographs. In this study, a popular commercial spectrophotometric serum cholesterol test kit was evaluated for the analysis of plant sterols. The results indicate that the method could be modified to analyze phytosterols and phytostanols by increasing the incubation time. Both free phytosterols and fatty acyl phytosteryl esters were quantitatively analyzed, but ferulate phytosteryl esters, such as those that are found in corn and other cereals, were not hydrolyzed by the enzymes in the test kit and therefore were not detected.

Previous attempts at separating nonpolar lipid esters (including wax esters, sterol esters, and methyl esters) have achieved only limited success. Among the several normal-phase methods tested, a single recent report of a method employing an alumina column at 30 degrees C with a binary gradient system was the most promising. In the current study, modification of the alumina method by increasing the column temperature to 75 degrees C improved the separation of standards of wax esters and sterol esters. Elevated column temperature also enhanced the separation of FAME with differing degrees of unsaturation. Evidence was also presented to indicate that the method similarly separated phytosterol esters, based on their levels of unsaturation. With the increased interest in phytosterol- and phytostanol-ester enriched functional foods, this method should provide a technique to characterize and compare these products.

Phytosterols (plant sterols) are triterpenes that are important structural components of plant membranes, and free phytosterols serve to stabilize phospholipid bilayers in plant cell membranes just as cholesterol does in animal cell membranes. Most phytosterols contain 28 or 29 carbons and one or two carbon-carbon double bonds, typically one in the sterol nucleus and sometimes a second in the alkyl side chain. Phytostanols are a fully-saturated subgroup of phytosterols (contain no double bonds). Phytostanols occur in trace levels in many plant species and they occur in high levels in tissues of only in a few cereal species. Phytosterols can be converted to phytostanols by chemical hydrogenation. More than 200 different types of phytosterols have been reported in plant species. In addition to the free form, phytosterols occur as four types of "conjugates," in which the 3beta-OH group is esterified to a fatty acid or a hydroxycinnamic acid, or glycosylated with a hexose (usually glucose) or a 6-fatty-acyl hexose. The most popular methods for phytosterol analysis involve hydrolysis of the esters (and sometimes the glycosides) and capillary GLC of the total phytosterols, either in the free form or as TMS or acetylated derivatives. Several alternative methods have been reported for analysis of free phytosterols and intact phytosteryl conjugates. Phytosterols and phytostanols have received much attention in the last five years because of their cholesterol-lowering properties. Early phytosterol-enriched products contained free phytosterols and relatively large dosages were required to significantly lower serum cholesterol. In the last several years two spreads, one containing phytostanyl fatty-acid esters and the other phytosteryl fatty-acid esters, have been commercialized and were shown to significantly lower serum cholesterol at dosages of 1-3 g per day. The popularity of these products has caused the medical and biochemical community to focus much attention on phytosterols and consequently research activity on phytosterols has increased dramatically.

Dry-milled yellow corn and freshly ground food and nonfood grade yellow and white hybrid corn kernels were pretreated in a solution of lactic acid and sodium metabisulfite followed by extraction with 70% ethanol. Zein was precipitated from the extract by reducing the ethanol content of the extract to 40%. Lipid associated with the zein isolates was between 15 and 20% and contained mostly endogenous free fatty acids. The effect of the endogenous free fatty acids on zein isolate films, with and without free fatty acids, was determined by measuring various film properties. Stress-strain measurements indicated 40-200% greater elongation for zein films containing endogenous free fatty acids. Films prepared from zein isolated from preground corn stored for approximately 4 months (27 degrees C, 17% relative humidity) had approximately 3 times greater elongation values than zein films prepared from freshly ground corn.

Phytosterols and phytostanols are known to lower low-density lipoprotein-cholesterol (LDL-C) levels in humans by up to 15%, and at least two products, Benecol and Take Control, are now on the market as naturally derived fatty acid esters of phytostanols (stanol esters) and phytosterols (sterol esters), respectively. A synthetic process was developed to synthesize gram quantities of trans-feruloyl-beta-sitostanol from ferulic acid and beta-sitostanol, with high purity and yields of approximately 60%. The process involves (a) condensation of trans-4-O-acetylferulic acid with the appropriate phytostanol or phytostanol mixture in the presence of N,N-dicyclohexylcarbodiimide and 4-(dimethylamino)pyridine, (b) separation of the trans-4-O-acetylferuloyl products by preparative liquid chromatography, (c) selective deacetylation of the feruloyl acetate, and (d) chromatographic purification of the feruloylated phytostanols. The process was successfully applied to synthesize stanol trans-feruloyl esters from "Vegetable Stanols", a mixture of approximately 70:30 beta-sitostanol and beta-campestanol, in comparable purity and yield.

Seeds of 49 accessions of corn (Zea mays ssp. mays), 9 accessions of teosinte (Zea species that are thought to be ancestors and probable progenitors to corn), and 3 accessions of Job's tears (Coix lacryma), obtained from a germplasm repository, were ground and extracted with hexane. Whole kernel oil yields and levels of four phytonutrients (free phytosterols, fatty acyl phytosterol esters, ferulate phytosterol esters, and gamma-tocopherol) in the oils were measured. Among the seeds tested, oil yields ranged from 2.19 to 4.83 wt %, the levels of ferulate phytosterol esters in the oil ranged from 0.047 to 0.839 wt %, the levels of free phytosterols in the oil ranged from 0.54 to 1.28 wt %, the levels of phytosterol fatty acyl esters in the oil ranged from 0.76 to 3.09 wt %, the levels of total phytosterols in the oil ranged from 1.40 to 4.38 wt %, and the levels of gamma-tocopherol in the oil ranged from 0.023 to 0.127 wt %. In general, higher levels of all three phytosterol classes were observed in seed oils from accessions of Zea mays ssp. mays than in seed oils from accessions of the other taxonomic groups. The highest levels of gamma-tocopherol were observed in teosinte accessions.

Extraction of corn bran or corn fiber with polar solvents such as methylene chloride, ethanol or chloroform/methanol yielded common lipids and two unknown high-performance liquid chromatography (HPLC) peaks, each with an ultraviolet absorbance maximum at 320 nm. HPLC-mass spectrometry revealed that the unknowns were diferuloylputrescine (DFP) and p-coumaroyl-feruloylputrescine (CFP). When compared to extracts of corn fiber (a pericarp-enriched fraction from the wet milling of corn), comparable extracts of corn bran (a pericarp- enriched fraction from the dry milling of corn) yielded three- to eightfold higher levels of DFP and CFP. Extraction of corn bran or fiber with an accelerated solvent extractor revealed that elevated temperatures greatly enhanced the extraction of DFP and CFP by methylene chloride and ethanol. "Corn bran oil," prepared by extraction of corn bran with hot methylene chloride, contained 14 wt% DFP and 3 wt% CFP. However, when hexane was used as a solvent, accelerated solvent extraction of the corn bran or fiber did not extract any DFP or CFP. Extraction of wheat bran or psyllium hulls with hot methylene chloride did not yield any detectable DFP or CFP. Because it has been suggested that polyamine conjugates such as DFP and CFP may function as natural pesticides, a rapid method was developed to purify them so that their biological activity could be evaluated.

Corn hulls are composed of two major layers: the outer layer, the pericarp, is made up of non-living cell walls, and an inner layer, the aleurone, consists of a single layer of living cells, surrounded by thick cell walls. Dissected pure pericarp and aleurone fractions were ground and extracted with hexane and the yields and compositions of the resulting oils were examined. This study revealed that the high levels of ferulate-phytosterol esters and the high concentration of sitostanol previously reported in corn-fibre oil actually originate in the aleurone cells.

Thirty-eight strains of filamentous bacteria, many of which are thermophilic or thermotolerant and commonly found in composts and mouldy fodders, were examined for their ability to produce cutinolytic esterase (cutinase) in culture media supplemented with cutin, suberin or cutin-containing agricultural by-products. Initially, the ability of culture supernatants to hydrolyse the artificial substrate p-nitrophenyl butyrate was determined by spectrophotometric assays. Only one bacterium, Thermoactinomyces vulgaris NRRL B-16117, exhibited cutinolytic esterase production. The enzyme was highly inducible, was repressed by the presence of glucose in the medium and hydrolysed both apple and tomato cutins. Inducers included apple cutin, apple pomace, tomato peel, potato suberin and commercial cork. Unlike similar fungal enzymes, the T. vulgaris cutinolytic esterase was not inducible by cutin hydrolysate. The cutinolytic esterase exhibited a half-life of over 60 min at 70 degrees C and a pH optimum of >/= 11.0. This study indicates that thermophylic filamentous bacteria may be excellent commercial sources of heat-stable cutin-degrading enzymes that can be produced by fermentation of low cost feedstocks.

An inexpensive zein-lipid mixture was isolated from yellow dent, dry-milled corn. Grease permeation through zein isolate applied to brown Kraft paper was found to be independent of loading levels at zein isolate levels above 30 mg/16 in.(2). The data shows that water vapor transmission rates depended on the amount of coating applied. Triacylglycerols were the most abundant lipid in milled corn but were absent in the zein isolate (perhaps due to hydrolysis by lipases). Zein from the paper was hydrolyzed enzymatically and the hydrolysis monitored by SDS-capillary electrophoresis. At an E:S ratio of 1:100 no further increase in the hydrolysate peak occurred after 10 and 30 min for alpha-chymotrypsin and pancreatin 8 x; however, zein and lipid were still present 1 h after hydrolysis by pancreatin 1 x.

Previously, hexane extraction of corn fiber was reported to produce a unique and potentially valuable oil that contained high levels of several phytosterols (which have been noted for their cholesterol-lowering properties). Current studies revealed that heat treatment (over the range of 100-175 degrees C) of corn fiber in either a convection oven or a vacuum oven caused only a modest reduction in the levels of the phytosterol components. However, these same heat pretreatments caused a considerable increase (up to 10-fold) in the levels (increasing from 0.34 wt % to a maximum of 3.64 wt % gamma-tocopherol in the oil) and yields (increasing from 5.4 mg of gamma-tocopherol/100 g of corn fiber to a maximum of 52.1 mg of gamma-tocopherol/100 g of corn fiber) of gamma-tocopherol in corn fiber oil. The main differences between the convection oven and vacuum oven pretreatments were associated with the disappearance of free fatty acids and free phytosterols at the higher temperature pretreatments in the vacuum oven, probably due to the lower boiling points of these lipids. Microwave pretreatment was also effective but caused a much smaller increase in the levels of gamma-tocopherol.

The tapetum cells in the developing anthers of Brassica napus contained abundant elaioplasts, which had few thylakoid membranes but were packed with globuli of neutral esters. Of the neutral esters, the major ester group possessed mainly 24-methylenecholesterol, 31-norcycloartenol, 24-dehydropollinastanol, and pollinastanol esterified to 18:3 and other unsaturated and saturated fatty-acyl moieties. The minor ester group had a dominant component tentatively identified as 12-dehydrolupeol esterified to mostly 18:0, 16:0, and 20:0 fatty-acyl moieties. The elaioplasts also contained a high proportion (16% w/w of total lipids) of monogalactosyldiacylglycerols (MGDG). This is the first report of plastids having steryl esters as the predominant lipids. We propose that the globuli contain steryl esters and are stabilized by surface MGDG and structural proteins. The tapetosomes, the other abundant lipid-containing organelles in the tapetum, possessed triacylglycerols (TAG) as the predominant lipids. At a late stage of anther development, the minor group of neutral esters and MGDG of the elaioplasts, as well as the TAG of the tapetosomes, were degraded. Steryl esters similar to those of the elaioplasts were recovered from the pollen surface and were the major lipids of the pollen coat. The pollen coat steryl esters and proteins could be extracted with moderately polar or nonpolar solvents. These proteins, which were mostly fragments of oleosins derived from the tapetosomes, had a high proportion of lysine (13 mol %). The possible functions of the steryl esters and the proteins on the pollen surface are discussed.

Cellular lipids were extracted from three species of Oomycete plant pathogens (Pythium ultimum, Phytophthora infestans, and Ph. capsici) and analyzed via normal-phase high-performance liquid chromatography with flame-ionization detection. The most abundant polar lipids in each of the three species were the polar membrane lipids, phosphatidylethanolamine (PE), phosphatidylcholine, and a phosphosphingolipid that eluted soon after PE. Structural analysis via mass spectrometry and nuclear magnetic resonance spectrometry revealed that the phosphosphingolipid was ceramide phosphorylethanolamine (Cer-PE). The most abundant molecular species of Cer-PE in P. ultimum had a molecular weight of 670.5, contained an unusual 19-carbon branched triunsaturated sphingoid (C19-delta 4, 8, 10, 9-methyl long-chain base) and palmitic acid as the amide-linked fatty acid. The most abundant molecular species of Cer-PE in Ph. infestans had a molecular weight of 714.5, contained a common 16-carbon 1,3 di-OH sphingoid, and erucic (cis 13-docosenoic, C22-delta 13) acid as the amide-linked fatty acid. The Cer-PE in Ph. capsici comprised a mixture of each of the two molecular species found in P. ultimum and Ph. infestans.

Zymomonas mobilis (ATCC 29191) was grown either aerobically or anaerobically in the presence of 2% (wt/vol) glucose and 0, 3, or 6% (vol/vol) ethanol. The rates of growth and the composition of hopanoids, cellular fatty acids, and other lipids in the bacterial membranes were quantitatively analyzed. The bacterium grew in the presence of 3% and 6% ethanol and was more ethanol tolerant when grown anaerobically. In the absence of ethanol, hopanoids comprised about 30% (by mass) of the total cellular lipids. Addition of ethanol to the media caused complex changes in the levels of hopanoids and other lipids. However, there was not a significant increase in any of the hopanoid lipid classes as ethanol concentration was increased. As previously reported, vaccenic acid was the most abundant fatty acid in the lipids of Z. mobilis, and its high constitutive levels were unaffected by the variations in ethanol and oxygen concentrations. A cyclopropane fatty acid accounted for 2.6-6.4 wt % of the total fatty acids in all treatments.

Tetrahydroxybacteriohopane (1), a bacterial hopanoid, inhibited soybean 15-lipoxygenase with an IC50 of about 10 microM. After per-O-acetylation of 1 no inhibition of the 15-lipoxygenase was observed. Two other bacterial hopanoids, tetrahydroxybacteriohopane glucosamine (2) and tetrahydroxybacteriohopane ether (3), stimulated the activity of soybean 15-lipoxygenase. The activities of two other arachidonic acid-metabolizing enzymes, human 5-lipoxygenase and prostaglandin H synthase, were unaffected by 1.

The acyl binding site of Rhizopus delemar prolipase and mature lipase was altered through site-directed mutagenesis to improve lipase specificity for short- or medium-chain length fatty acids. Computer-generated structural models of R. delemar lipase were used in mutant protein design and in the interpretation of the catalytic properties of the resulting recombinant enzymes. Molecular dynamics simulations of the double mutant, val209trp + phe112trp, predicted that the introduction of trp112 and trp209 in the acyl binding groove would sterically hinder the docking of fatty acids longer than butyric acid. Assayed against a mixture of triacylglycerol substrates, the val209trp + phe112trp mature lipase mutant showed an 80-fold increase in the hydrolysis of tributyrin relative to the hydrolysis of tricaprylin while no triolein hydrolysis was detected. By comparison, the val94Trp mutant, predicted to pose steric or geometric constraints for docking fatty acids longer than caprylic acid in the acyl binding groove, resulted in a modest 1.4-fold increase in tricaprylin hydrolysis relative to the hydrolysis of tributyrin. Molecular models of the double mutant phe95asp + phe214arg indicated the creation of a salt bridge between asp95 and arg214 across the distal end of the acyl binding groove. When challenged with a mixture of triacylglycerols, the phe95asp + phe214arg substitutions resulted in an enzyme with 3-fold enhanced relative activity for tricaprylin compared to triolein, suggesting that structural determinants for medium-chain length specificity may reside in the distal end of the acyl binding groove. Attempts to introduce a salt bridge within 8 A of the active site by the double mutation leu146lys + ser115asp destroyed catalytic activity entirely. Similarly, the substitution of polar Gln at the rim of the acyl binding groove for phe112 largely eliminated catalytic activity of the lipase.

Lipid compositional analysis was conducted on the white, yellow, and brown cyst stages of Globodera rostochiensis (golden cyst nematode). Triacylglycerols were the largest lipid fraction in all stages examined, ranging from 55-75% of total lipid. Ethanolamine phosphoglycerides and choline phosphoglycerides were present in high amounts in all cyst fractions, with a total phospholipid content of 20%, 14.7%, and 12.8% in the white, yellow, and brown cyst stages, respectively. Sterols, steryl esters, sphingomyelin, and cardiolipin were found in minor amounts in all three cyst stages and showed greater changes than other classes of lipids relative to cyst stage. The fatty acid compositions of the three cyst stages were similar. Eicosenoic acid (20:1) and arachidonic acid (20:4) were found in higher concentrations than other fatty acids in all cyst preparations; vaccenic acid (18:1) occurred at the third highest concentration. More than 78% of total fatty acids were unsaturated at all cyst stages, and more than 60% were of C20 or longer chain length. The lipid profile of all three cyst stages is consistent with invertebrate adaptation to low-temperature environments.

Hopanoids and other lipids were extracted from Zymomonas mobilis and quantitatively analyzed by high-performance liquid chromatography. Previous methods for hopanoid analysis required derivatization of the hopanoids via periodate oxidation or acetylation. The current method employs a normal-phase silica gel column, a ternary gradient of hexane-isopropanol-water-triethylamine, and detection with a flame ionization detector. Three major hopanoid classes were separated and quantified by this new method, and together they comprised 30 to 40% of the total lipid in these cells. Mass spectrometry confirmed that chemical structures of these hopanoids were consistent with those previously proposed. In addition, several other common lipid classes (free fatty acids and six classes of common phospholipids), comprising the remaining 60 to 70% of the total lipids, were also separated and quantified. This new chromatographic method represents the first technique for the analysis and purification of intact hopanoids. This method should be useful for the analysis and purification of hopanoids from other bacterial species.