Two-step enzymatic synthesis of docosahexaenoic acid-rich symmetrically structured triacylglycerols via 2-monoacylglycerols

Symmetrically structured triacylglycerols (TG) rich in docosahexaenoic acid (DHA) with caprylic acid (CA) at the outer positions were synthesized enzymatically form bonito oil in a two-step process: (i) ethanolysis of bonito oil TG to 2-monoacylglycerols (2-MG) and fatty acid ethyl esters, and (ii) reesterification of 2-MG with ethyl caprylate. Ethanolysis catalyzed by immobilized Candida antarctica lipase (Novozym 435) yielded 92.5% 2-MG with 43.5% DHA content in 2 h. The 2-MG formed were reesterified with ethyl caprylate by immobilized Rhizomucor miehei lipase (Lipozyme IM) to give structured TG with 44.9% DHA content [based on fatty acid composition with caprylic acid (CA) excluded] in 1 h. The final structured lipids comprised 85.3% TG with two CA residues and one original fatty acid residue, 13% TG with one CA residue and two original fatty acid residues, and 1.7% tricaprylolglycerol (weight percent). The amount of TG with two CA residues and one C₂₂ residue (22∶6=DHA, 22∶5, and 22∶4) was 51 wt%. The 1,3-dicapryloyl-2-docosahexaenoylglycerol to 1,2(2,3)-dicapryloyl-3 (1)-docosahexaenoylglycerol ratio (based on high-performance liquid chromatography peak area percentages) was greater than 50∶1. The recovery of TG as structured lipids after silica gel column purification was approximately 71%. Ethyl esters and 2-MG formed at 2 h of ethanolysis could be used to determine the positional distribution of fatty acids in the intial TG owing to the high 1,3-regiospecificity of Novozym 435 and the reduced acyl migration in the system.     

Inhibition of growth of Escherichia coli O157:H7 in fresh radish (Raphanus sativus L.) sprout production by calcinated calcium

The inhibitory effect of calcinated calcium on the growth of Escherichia coli O157:H7 during fresh radish (Raphanus sativus L.) sprout production was studied. It was revealed that the addition of 0.4% (wt/vol) calcinated calcium into radish sprouting medium which was artificially contaminated with E. coli O157:H7 (3.0 to 3.2 log CFU/ml) completely inhibited the growth or inactivated the microorganism. When radish seed extract was used instead of radish sprout production, the same extent of growth inhibition or inactivation was observed with much lower amounts (0.07%) of calcinated calcium under similar experimental conditions. The findings suggested that calcinated calcium may be useful to control E. coli O157:H7 contamination during the production of radish sprouts.     

Increasing n-3 polyunsaturated fatty acid content of fish oil by temperature control of lipase-catalyzed acidolysis

An attempt was made to further increase the content of n-3 polyunsaturated fatty acid (n-3 PUFA) of fish oil by lipase-catalyzed acidolysis (reaction between fish oil and n-3 PUFA-enriched free fatty acid) without solvent. A bioreactor system was constructed composed of a water-jacketed packed-bed column and a substrate reservoir with a circulation pipeline between the packed-bed column and the reservoir. By keeping the temperature of the reservoir at −10°C (for the first 20 h), followed by −20°C (for the subsequent 40 h) during the batch acidolysis, crystals of free fatty acid appeared, which were removed intermittently by a cotton plug packed in the tip of the outlet pipe in the reservoir. The n-3 PUFA content of the triacylglycerol fraction increased a further 10% by the reduced temperature of the reservoir. Bioreactors for Enzymatic Reaction of Fats and Fatty Acid derivatives, Part XV.     

Evaluation of protein adsorption on hydrogenated amorphous carbon films by surface plasmon resonance phenomenon

To develop an anti-thrombogenic coating, hydrogenated amorphous carbon (a-C:H) and related films were studied in terms of their protein adsorption during the initial process in thrombogenesis by surface plasmon resonance (SPR) phenomenon using a multilayer device consisting of an a-C:H layer on Au. Two a-C:H films with different hydrogen contents, a nitrogenated a-C:H (a-C:N:H) and a fluorinated amorphous carbon (a-C:F) film were prepared on the Au layer in the multilayer device. Human serum albumin (HSA) in a phosphor buffer (PB) was used as a protein. Na₂HPO₄·12H₂O, NaH₂PO₄·2H₂O and deionized water were mixed to coordinate PB. From the attenuation of reflected light, the SPR angle was determined to the angle at minimum reflection intensity. The observed behavior of the SPR angle indicated that HSA was adsorbed on all films. The SPR angle was analyzed to estimate the multilayer index of the HSA-adsorbed layer on each film. The HSA adsorption ability of both a-C:H and a-C:N:H films was similar, and the absorption ability of the a-C:F film was lower than that of the other films. Hence, the surface polarization dominates the adsorption ability of HSA on a-C:H films and related film.     

High-yield diacylglycerol formation by solid-phase enzymatic glycerolysis of hydrogenated beef tallow

Diglyceride (DG) was prepared by reaction of hydrogenated beef tallow and glycerol in the presence of aPsesudomonas lipase. The yield of DG depended strongly on the reaction temperature. After initial incubation at 60°C for 2 h, followed by the first temperature shift down to 55°C for 4 h and then the second shift down to 48°C for up to 3 d, the reaction mixture became solid and a yield of approximately 90% DG was obtained. About 95% of total DG was 1,3-DG. The yield of DG was also dependent on the glycerol (GL) to triglyceride (TG) molar ratio. At the molar ratio of 1∶2 (GL/TG), the enzyme-catalyzed reaction was highly efficient and utilized essentially all of the glycerol. The free fatty acid (FFA) content at equilibrium depended on the water concentration in the glycerol phase. The initial rate of FFA formation was low and was hardly affected by the moisture content between 0.5 and 4%, but, at higher water content (4–6.7%), there was a small increase in the rate.     

New Sweet Potato Line having low Gelatinization Temperature and altered Starch Structure

A new sweet potato breeding line, Kanto 116, was developed, featuring low gelatinization temperature and an altered starch fine structure. Starch granules from Kanto 116 showed an abnormal morphology characterized by cracking into granules. Starch content, amylose content and tuberous root appearance of Kanto 116 were similar to those of the control and the parents. Pasting temperatures of Kanto 116 starch determined by the Rapid Visco Analyser were 51.4 — 52.6 °C, approximately 20 °C lower than those of the control and parents starches. Onset, peak, and conclusion temperature of gelatinization, and gelatinization enthalpy of Kanto 116 starch determined by differential scanning calorimetry were 39.0 °C, 46.9 °C, 64.8 °C, and 8.8 J/g, respectively, and much lower than those of the control and parents starches. The chain‐length distribution of the amylopectin molecules, determined by high‐performance anion‐exchange chromatography, showed that Kanto 116 starch had a higher proportion of short chains (DP 6 — 11) and a lower proportion of chains between DP 12 — 28 than control and parent starches. The debranched β‐limit dextrin of Kanto 116 starch also showed that the proportion of both short and long B1 chains was different from those of the control and parents starches.