Alan R. McCurdy

Detachment ofPseudomonas fluorescens from biofilms on glass surfaces in response to nutrient stress

The effects of glucose and nitrogen depletion on the colonization of glass Petri plates byPseudomonas fluorescens were studied in batch culture. Colonization of the surfaces was initiated before colonization of the bulk phase, and biofilm formation was observed. This resulted in an apparent lag in the batch growth curve for the cell suspension. The lag phase was an artifact caused by the partitioning of cells between the bulk and solid phase of the culture and was not due to a reduction in the growth rate of unattached cells. The specific growth rate of the unattached cells (0.331 hour−1) was almost twice that determined for the total population (0.171 hour−1). Consequently the growth rate of biofilm-forming bacteria cannot be determined in batch culture unless the growth of both attached and unattached cells is monitored, and batch growth curves may contain artifacts due to the formation and dispersion of biofilms. The depletion of either glucose or nitrogen led to the active detachment of cells from the biofilm. An increase in the hydrophobicity of unattached cells was noted on depletion of carbon. This increase was the result of emigration of cells from the surface into the bulk phase.

Enzymatic pretreatment of deodorizer distillate for concentration of sterols and tocopherols

Separation of sterols and tocopherols from fatty acids in deodorizer distillate was facilitated through lipase-catalyzed modification of fatty acids in canola, mixed and soya deodorizer distillates. The fatty acid esterification with methanol catalyzed by SP-382 (an immobilized nonspecific lipase) proceeded rapidly, with conversion of fatty acid to methyl ester in 5 h being 96.5, 83.5 and 89.4%, respectively. A model mixture of pure oleic acid and dl-α-tocopherol was used to study any potential side reactions that may lower the tocopherol content during the esterification reaction. Under the conditions employed, the loss of tocopherol was less than 5%. Simple vacuum distillation (1–2 mm Hg) was employed to remove the volatile fraction (methyl esters of fatty acids, some fatty acids and other volatiles) of the esterified deodorizer distillate, leaving behind sterols, sterol esters and tocopherols. Sterols and tocopherols were almost completely retained in the residue fraction with recoveries in the range of 95%. Overall recoveries of sterols and tocopherols after esterification and distillation were over 90% for all the deodorizer distillate samples.

Lipase-catalyzed esterification of oleic acid and methanol in hexane—A kinetic study

The kinetics of immobilized lipase-catalyzed esterification of oleic acid and methanol in hexane were investigated. The reaction follows Michaelis-Menton kinetics as observed from the relationship of initial rate of the reaction, both as a function of enzyme and of substrate concentration. Inhibition by excess of methanol has been identified. The kinetic constants have been measured for the reaction in the absence of any significant external diffusional limitations. The kinetics of the enzymatic reaction are suggested to agree with a Ping-Pong Bi Bi mechanism.

Detection of olive oil adulteration with canola oil from triacylglycerol analysis by reversed-phase high-performance liquid chromatography

The objective of this study was to explore the use of reversed-phase high-performance liquid chromatography (RP-HPLC) as a means to detect adulteration of olive oil with less expensive canola oil. Previously this method has been shown to be useful in the detection of some other added seed oils; however, the detection of adulteration with canola oil might be more difficult due to similarities in fatty acid composition between canola oil and olive oil. Various mixtures of canola oil with olive oils were prepared, and RP-HPLC profiles were obtained. Adulteration of olive oil samples with less than 7.5% (w/w) canola oil could not be detected.

Stereospecific analyses of seed triacylglycerols from high-erucic acid brassicaceae: Detection of erucic acid at thesn-2 position inBrassica oleracea L. Genotypes

Stereospecific analyses of triacylglycerols from selected high-erucic acid breeding lines or cultivars ofBrassica napus L. andB. oleracea L. have been performed. Initial lipase screening revealed that while allB. napus lines contained little or no erucic acid at thesn-2 position, several of theB. oleracea lines had significant proportions of erucic acid at this position. Detailed stereospecific analyses were performed on the triacylglycerols from these lines by using a Grignard-based deacylation, conversion of thesn-1,sn-2 andsn-3 monoacylglycerols to their di-dinitrophenyl urethane (DNPU) derivatives, resolution of the di-DNPU-monoacylglycerols (MAGs) by high-performance liquid chromatography on a chiral column, transmethylation of eachsn-di-DNPU MAG fraction and analysis of the resulting fatty acid methyl esters by gas chromatography. The findings unequivocally demonstrate for the first time that, within the Brassicaceae, there existsB. oleracea germplasm containing seed oils with substantial erucic acid (30–35 mol%) at thesn-2 position. This has important implications for biotechnology and breeding efforts designed to increase the levels of erucic acid in rapeseed beyond 66 mol% to supply strategic industrial feedstocks. In the first instance, the germplasm will be of direct use in retrieving a gene encoding aBrassica lyso-phosphatidic acid acyltransferase with an affinity for erucoyl-CoA. In a breeding program, the germplasm offers promise for the introduction of this trait intoB. napus by interspecific hybridization and embryo rescue.

Effect of randomization on oxidative stability of vegetable oils at two different temperatures

Five vegetable oils were each randomized by chemical means (with the use of a sodium potassium alloy) and by enzymatic means (using a nonspecific lipase). The success of the randomization procedure was confirmed via positional analysis. The oxidative stabilities of the native and chemically randomized oils were determined at storage temperatures of 28°C and 55°C using absorbance at 234 nm as indicative of conjugated diene content. No difference between curves occurred in oils stored at 55°C, however, at the lower temperature all chemically randomized oils had significantly steeper slopes (P<0.05), suggesting a lower stability. When both enzymatically and chemically randomized oils were compared to native oils at 28°C, no significant difference occurred between slopes of native and enzymatically randomized oils, however, the end content of conjugated dienes was significantly higher for chemically randomized canola, corn and soybean oils (P<0.05). No difference was seen between the slopes of the three different oils from either linseed or sunflower. Since both of these oils exhibited higher oxidation rates, it is possible that observation of differences between the stability of native and chemically randomized oild is dependent upon the rate of the reaction.

Thermal resistance of Streptococci isolated from pasteurized ham

Abstract Streptococcus faecium and Streptococcus faecalis, two examples of thermoduric spoilage organisms from perishable cured meats, were isolated from pasteurized canned hams and tested for heat resistance using the multiple point method. Known cell concentrations were inoculated into two tempered liquid menstrua (Soren-sons buffer and a derived ham broth), and exposed to a variety of heating temperatures. Survivor curves, phantom thermal death time curves, D values, and z values were determined. Time-temperature relationships of interest to industry were obtained, and it was found that Streptococcus faecium was more heat resistant than Streptococcus faecalis. Four of the six organisms tested were more resistant in the ham menstrum than in the buffer.

Enzymatic methylation of canola oil deodorizer distillate

Methylation of canola oil deodorizer distillate catalyzed by a nonspecific lipase was investigated. The conversion of fatty acids to methyl esters has been optimized by using a statistical design. Up to 96.5% conversion of fatty acids to their methyl esters has been achieved without the aid of vacuum or any water-removing agent. The effects of temperature, ratio of the reactants (methanol: fatty acids in the deodorizer distillate) and enzyme concentration on the equilibrium conversion were studied. The temperature and ratio of the reactants showed a significant effect on the conversion of fatty acids to methyl esters and they exhibited a strong interactive effect. Enzyme concentration in the range of 2.7% to 4.3% did not show a significant effect on the equilibrium conversion of fatty acids. Greater than 95% conversion of fatty acids to methyl esters was achieved at temperatures around 50°C and at a ratio of the reactants between 1.8 and 2.0. The inhibitory effect of hydrophilic methanol on the enzyme activity was largely reduced by working at the lower temperature range (around 50°C).

Application of reversed-phase liquid chromatography and prepacked C18 cartridges for the analysis of oxytetracycline and related compounds

The reversed-phase (RP) chromatographic separation of oxytetracycline (OTC) 4-epioxytetracycline (4-epiOTC), alpha-apooxytetracycline (alpha-apoOTC), and beta-apooxytetracycline (beta-apoOTC) has been accomplished on an Inertsil C8 column at ambient temperature. Using the simplex method of solvent optimization, a 0.1 M ammonium acetate buffer (pH 3.0)-acetonitrile-tetrahydrofuran (72.5:12.5:15, v/v/v) mobile phase was found to give excellent separation of the compounds. OTC, 4-epiOTC, alpha-apoOTC and beta-apoOTC were resolved in 35 min with calculated detection limits of 40, 20, 50 and 140 ng/ml, respectively. Solid-phase extraction (using RP C18 cartridges) of OTC and OTC degradation compounds from distilled water and porcine muscle was tested at four concentration levels ranging from 200 to 2000 ng/ml (g); overall mean recovery of OTC from distilled water and porcine tissue was greater than 90% and 70%, respectively.

The effect of randomization on the stability of blends of trioleoylglycerol and linseed oil

The effect of fatty acid arrangement on triacylglycerols was determined by assessing the stability of mixtures of trioleoylglycerol (consisting of a comparatively stable fatty acid) and linseed oil (containing a high amount of unstable fatty acids in the form of linoleic and linolenic acids) in the ratios of 90:10, 85:15, 80:20, 70:30 and 60:40 (w/w), respectively, before and after enzymatic randomization. Randomization resulted in increased stability; however, increasing the content of the unstable triacylglycerol resulted in a decrease in this effect. Based on these results, it was concluded that randomization of triacylglycerols can have a positive effect on the oxidative stability of an oil if the content of autoxidatively unstable triacylglycerols is low in the original blend. This results in substantial dilution of unstable fatty acids among the more stable triacylglycerols upon randomization.