24-h Hydrolysis Research Articles

Pretreatment of Jerusalem artichoke stalk using hydroxylammonium ionic liquids and their influences on 2,3-butanediol fermentation by Bacillus subtilis

Pretreatment of lignocellulose is a vital step for biological production of bio-chemicals and bio-fuels. In this work, the pretreatment of Jerusalem artichoke stalk (JAS) by hydroxylammonium ionic liquids was evaluated based on pretreatment efficiency including polysaccharide recovery and enzymatic digestibility, and influence of ionic liquids on 2,3-butanediol fermentation using Bacillus subtilis. The results showed ethanolammonium acetate (EOAA) was efficient in JAS pretreatment, and maximum cell density was increased 25% when EOAA concentration was not greater than 0.3 mol/L in medium, while the total concentration of acetoin and 2,3-butanediol was 15% greater than the control at 0.1 mol/L EOAA. After the pretreatment under optimized conditions of 170 °C for 5-h and liquid–solid ratio of 18, about 87% cellulose and 75% hemicellulose were recovered, and glucose yield of 64% and xylose of 66% were obtained after 24-h hydrolysis of JAS residue by cellulase (15 FPU/g) with solid loading of 10 wt%.

Statistical analysis of the crystallinity index of nanocellulose produced from Kraft pulp via controlled enzymatic hydrolysis.

Enzymatic hydrolysis processes can change the physical characteristics of nanocellulose derived from Kraft pulp. Among these attributes are its crystallinity index and dimensions. In this study, we determined the optimal conditions under which nanocellulose could be produced enzymatically with the greatest increase of the crystallinity index relative to its initial state. Application of Central Composite Rotatable Design statistical analysis to the experiments was employed to direct an increase the crystallinity index in 10% at the 24-H hydrolysis time. Upon establishment of ideal levels of starting material and enzyme, reactions were carried out at hydrolysis times of 24, 48, and 72H under these ideal parameters. The effectiveness of deagglomeration was demonstrated by measuring the hydrodynamic diameter of the particles by dynamic light scattering. Scanning electron microscopy was performed on four samples, the original material, kraft pulp, and hydrolyzed biomaterials at 72H in the ideal parameters. The hydrolyzed material with the best statistical data, revealing a fiber diameter of 180nm, disclosing to be biomaterial with nanocellulose dimensions.

Predictions of ruminal outflow of essential amino acids in dairy cattle

The objective of this work was to update and evaluate predictions of essential AA (EAA) outflows from the rumen. The model was constructed based on previously derived equations for rumen-undegradable (RUP), microbial (MiCP), and endogenous (EndCP) protein outflows from the rumen, and revised estimates of ingredient composition and EAA composition of the protein fractions. Corrections were adopted to account for incomplete recovery of EAA during 24-h acid hydrolysis. The predicted ruminal protein and EAA outflows were evaluated against a data set of observed values from the literature. Initial evaluations indicated a minor mean bias for non-ammonia, non-microbial nitrogen flow ([RUP + EndCP]/6.25) of 16 g of N per day. Root mean squared errors (RMSE) of EAA predictions ranged from 26.8 to 40.6% of observed mean values. Concordance correlation coefficients (CCC) of EAA predictions ranged from 0.34 to 0.55. Except for Leu, all ruminal EAA outflows were overpredicted by 3.0 to 32 g/d. In addition, small but significant slope biases were present for Arg [2.2% mean squared error (MSE)] and Lys (3.2% MSE). The overpredictions may suggest that the mean recovery of AA from acid hydrolysis across laboratories was less than estimates encompassed in the recovery factors. To test this hypothesis, several regression approaches were undertaken to identify potential causes of the bias. These included regressions of (1) residual errors for predicted EAA flows on each of the 3 protein-driven EA flows, (2) observed EAA flows on each protein-driven EAA flow, including an intercept, (3) observed EAA flows on the protein-driven EAA flows, excluding an intercept term, and (4) observed EAA flows on RUP and MiCP. However, these equations were deemed unsatisfactory for bias adjustment, as they generated biologically unfeasible predictions for some entities. Future work should focus on identifying the cause of the observed prediction bias.

Estimation of correction factors to determine the true amino acid concentration of protein after a 24-hour hydrolysis

Although it has been acknowledged for a long time that a single period of hydrolysis, normally 21 to 24 h, is not the optimal time for most of the AA, a single period is routinely used due to time and cost constraints. As models to balance dairy rations for proteins are evolving toward balancing for AA, it becomes critical to improve the predictions of AA supply from digested proteins. Our objective was to develop correction factors that could systematically be applied to AA concentrations obtained after a 24-h hydrolysis of proteins to account for incomplete recovery and therefore determine their true AA composition. Thirteen substrates were selected to represent different types of proteins commonly used to estimate the supply of AA in ration formulation models: feed ingredients (grass silage, corn silage, soybean meal, canola meal, high-protein corn dried distillers grains, and wheat dried distillers grains plus solubles), 16-h rumen residues (soybean meal and canola meal), digesta (duodenal digesta and feces), and rumen microorganisms (fluid-associated bacteria, particle-associated bacteria and protozoa). Each protein was hydrolyzed in 6 N HCl for multiple hydrolysis times: 13 (2, 4, 8, 12, 18, 21, 24, 30, 48, 72, 96, 120, and 168 h) for feed ingredients, rumen residues, and digesta, and 9 (2, 4, 8, 18, 24, 30, 48, 96, and 168 h) for rumen microorganisms; all analyses were conducted in triplicate. Using nonlinear regression, the AA composition in the protein before the hydrolysis (A0) was derived for each AA in each protein. Two ratios were calculated as potential correction factors: A0/24-h concentration (A0/24h) and the maximal concentration/24-h concentration (max/24h). Both ratios were tested to determine if the type of proteins was affecting them. The ratios A0/24h were not affected by the type of proteins, whereas the ratios max/24h were also not affected by the type of proteins except for 3 nonessential AA (Ala, Glu, and Gly). In an attempt to propose correction factors, our results were combined with results from the literature reporting ratios A0/24h, ratios max/24h, or the ratio of the AA composition calculated from gene structure/24 h. The correction factors proposed for individual AA varied from 1.02 (Asp) to 1.12 (Thr). For the essential AA, the highest ratios were obtained, as expected, for the branched-chain AA and Thr. Formulation programs balancing dairy rations for essential AA would need to acknowledge the incomplete recovery of AA when obtained from 24-h hydrolysis and include correction factors, specific for each AA, but the same across different types of proteins, to correctly estimate the true AA supply to dairy cows.

Physico-chemical oxidative cleavage strategy facilitates the degradation of recalcitrant crystalline cellulose by cellulases hydrolysis

BackgroundEfficient enzymatic conversion of recalcitrant crystalline cellulose is critical for enabling cost-effective industrial conversion of cellulosic biomass to biofuels and chemicals. Fully understanding enzyme digestion mechanism is paving a new way to design efficient process for biomass conversion. Accordingly, a continuing drive is inspiring to discover new routes to promote crystalline cellulose disruption.ResultsHerein, a physico-chemical oxidative cleavage strategy of irradiation oxidation/post-reduction (IOPR) was employed to treat crystalline cellulose I to cleave glycosidic bonds association with some new oxidized and reduced chain ends, thus boosting downstream degradation by cellulases from Trichoderma reesei. The hydrolysis performance of treated crystalline cellulose was conducted with either T. reesei Cel7A (TrCel7A) alone, or a cellulase enzyme mixture (90% Celluclast 1.5 L, 10% β-glucosidase). 81.6 and/or 97% of conversion efficiency have been reached for 24-h and 48-h cellulase hydrolysis, respectively. The high efficient conversion of crystalline cellulose after IOPR is mainly attributed to generating some new chain ends, which are identified by MAIDI-TOF–MS and HPLC. Furthermore, the nanoscale architectures of crystalline cellulose before and after IOPR are systematically investigated by XRD, EPR, ATR- FTIR, GPC, and XPS techniques. Together with TEM images, the results reveal a fascinating digestion mechanism of “peel-off” and “cavity-formation” paradigms toward degrading crystalline cellulose by cellulase mixtures after IOPR treatment.ConclusionsThis encouraging results show that the proposed IOPR approach will become a potential competitive alternative to current biomass pretreatment. It opens a new avenue toward the implementation of pretreatment and the design of enzyme cocktails in lignocellulosic biorefinery.

Effects of reduced severity ammonia pretreatment on pelleted corn stover

Biomass densification impacts pretreatment efficacy and subsequent biochemical conversion to biofuels and other biobased chemicals. Pelleted corn stover was used to evaluate the soaking in aqueous ammonia (SAA) pretreatment efficacy at high solid loadings with reductions ammonia concentration, temperature, and pretreatment time. Pretreatment resulted in 70% greater delignification of stover pellets than loose stover. Glucose yields from enzymatic hydrolysis were 49% higher for the pelleted stover and yields did not change with a twofold increase in pretreatment solid loadings. Hydrolysis yields were modeled as a function of pretreatment conditions and the developed model predicted a maximum 24-h hydrolysis glucose yield of 96% at a pretreatment temperature of 60 °C, for 4 h, with 18% ammonia. Pretreatment severity can still be reduced while maintaining 90% or higher yields with different combinations of temperature, time, and ammonia concentration. Temperature was the most important pretreatment parameter within the design space for achieving high glucose yields. FTIR and SEM analysis of SAA-pretreated corn stover pellets illustrated that the pelleting process increases pretreatment efficacy by modifying the biomass structure, disrupting the lignin-hemicellulose linkages, and partially removing lignin. Using pelleted biomass can reduce the required severity of SAA-pretreatment while still producing glucose yields above 90%. Using pelleted corn stover as a biorefinery feedstock has potential to lower pretreatment costs in addition to the improved handling and transportation.

Durable and self-healing superhydrophobic polyvinylidene fluoride (PVDF) composite coating with in-situ gas compensation function

A durable and self-healing superhydrophobic polyvinylidene fluoride (PVDF)/polyester-modified silicone resin (PMSR)/microspheric Na2CO3@SiO2 coating with WCA of 158±1.5° and low SA of 4±0.5° has been successfully prepared via sol-gel and spraying technique. After 24h hydrolysis of tetraethylorthosilicate (TEOS), microspheric Na2CO3@SiO2 particles with extremely large storage spaces were obtained as reservoirs for the healing agent of POTS. The superhydrophobicity of the prepared coating can be recovered after 10 rubbing/healing test cycles, indicating the excellent self-healing ability. Simultaneously, the prepared coating also possesses gas compensation capability in acidic condition, which can extend superhydrophobic duration by 400%. The gas compensation capability can be mainly attributed to the bubbles generated from the reaction between acid and porous Na2CO3, hindering water wetting the surface. Moreover, the prepared coating can withstand 4217 friction cycles without obvious surface damage, which is approximately 6 times higher than that of pure PVDF coating. This is mainly due to the combined action of the high adhesion of PMSR, the self-lubrication of PVDF and the surface hierarchical nano/micro wrinkles. The prepared coating also shows prominent superhydrophobic stability under the conditions of acid/alkaline solution (pH=1–14) or strong UV exposure (14days). It is believed that this durable and self-healing superhydrophobic coating is anticipated to open a new avenue to enlarge the application range of superhydrophobic coatings.

Ruminal bacteria and protozoa composition, digestibility, and amino acid profile determined by multiple hydrolysis times.

Microbial samples from 4 independent experiments in lactating dairy cattle were obtained and analyzed for nutrient composition, AA digestibility, and AA profile after multiple hydrolysis times ranging from 2 to 168 h. Similar bacterial and protozoal isolation techniques were used for all isolations. Omasal bacteria and protozoa samples were analyzed for AA digestibility using a new in vitro technique. Multiple time point hydrolysis and least squares nonlinear regression were used to determine the AA content of omasal bacteria and protozoa, and equivalency comparisons were made against single time point hydrolysis. Formalin was used in 1 experiment, which negatively affected AA digestibility and likely limited the complete release of AA during acid hydrolysis. The mean AA digestibility was 87.8 and 81.6% for non-formalin-treated bacteria and protozoa, respectively. Preservation of microbe samples in formalin likely decreased recovery of several individual AA. Results from the multiple time point hydrolysis indicated that Ile, Val, and Met hydrolyzed at a slower rate compared with other essential AA. Singe time point hydrolysis was found to be nonequivalent to multiple time point hydrolysis when considering biologically important changes in estimated microbial AA profiles. Several AA, including Met, Ile, and Val, were underpredicted using AA determination after a single 24-h hydrolysis. Models for predicting postruminal supply of AA might need to consider potential bias present in postruminal AA flow literature when AA determinations are performed after single time point hydrolysis and when using formalin as a preservative for microbial samples.

Lignocellulosic butanol production from Napier grass using semi-simultaneous saccharification fermentation

Napier grass is a potential feedstock for biofuel production because of its strong adaptability and wide availability. Compositional analysis has been done on Napier grass which was collected from a local area of Taiwan. By comparing acid- and alkali-pretreatment, it was found that the alkali-pretreatment process is favorable for Napier grass. An overall glucose yield of 0.82g/g-glucosetotal can be obtained with the combination of alkali-pretreatment (2.5wt% NaOH, 8wt% sample loading, 121°C, and a reaction time of 40min) and enzymatic hydrolysis (40FPU/g-substrate). Semi-simultaneous saccharification fermentation (sSSF) was carried out, where enzymatic hydrolysis and ABE fermentation were operated in the same batch. It was found that after 24-h hydrolysis, followed by 96-h fermentation, the butanol and acetone concentrations reached 9.45 and 4.85g/L, respectively. The butanol yield reached 0.22g/g-sugarglucose+xylose. Finally, the efficiency of butanol production from Napier grass was calculated at 31%.

Assessment of oligogalacturonide from citrus pectin as a potential antibacterial agent against foodborne pathogens.

Foodborne diseases are an important public health problem in the world. The bacterial resistance against presently used antibiotics is becoming a public health issue; hence, the discovery of new antimicrobial agents from natural sources attracts a lot of attention. Antibacterial activities of oligogalacturonide from commercial microbial pectic enzyme (CPE) treated citrus pectin, which exhibits antioxidant and antitumor activities, against 4 foodborne pathogens including Salmonella Typhimurium, Staphylococcus aureus, Listeria monocytogenes, and Pseudomonas aeruginosa was assessed. Pectin hydrolysates from CPE hydrolysis exhibited antibacterial activities. However, no antibacterial activity of pectin was observed. Citrus oligogalacturonide from 24-h hydrolysis exhibited bactericidal effect against all selected foodborne pathogens and displayed minimal inhibitory concentration at 37.5 μg/mL for P. aeruginosa, L. monocytogenes, and S. Typhimurium, and at 150.0 μg/mL for S. aureus.

Physical, chemical and biochemical properties of casein hydrolyzed by three proteases: Partial characterizations

Sodium caseinate (NaCas) was hydrolyzed by papain, pancreatin and trypsin from 10min to 24h, and the hydrolysates were partially characterized for several important properties. At the studied conditions, papain and trypsin were more effective in hydrolyzing NaCas than pancreatin. Pancreatin treatments showed an initial increase in surface hydrophobicity, contrasting with the consistent decrease for the other two treatments. The solubility of NaCas at acidic pH was improved, becoming pH-independent after 24h hydrolysis. The emulsifying properties generally showed improvements after hydrolysis. The DPPH free radical scavenging activity, reducing power, and inhibition of linoleic acid autoxidation were significantly enhanced after appropriate hydrolysis, while metal ion chelating effects were slightly attenuated. The angiotensin converting enzyme-inhibitory activity was significantly improved by up to 9times than that of NaCas. These findings indicate that physical, chemical and biochemical properties of casein hydrolysates can be improved by selecting proteolytic conditions to produce functional ingredients.

Enzymatic pretreatment of lignocellulosic wastes to improve biogas production

The effect of enzymatic pretreatment of sugar beet pulp and spent hops prior to methane fermentation was determined in this study. These industrial residues were subjected to enzymatic digestion before anaerobic fermentation because of high fiber content (of 85.1% dry matter (DM) and 57.7% DM in sugar beet pulp and spent hops, respectively). Their 24h hydrolysis with a mix of enzymatic preparations Celustar XL and Agropect pomace (3:1, v/v), with endoglucanase, xylanase and pectinase activities, was most effective. Reducing sugars concentrations in hydrolysates of sugar beet pulp and spent hops were by 88.9% and 59.4% higher compared to undigested materials. The highest yield of biogas was obtained from the enzymatic hydrolysate of sugar beet pulp (183.39mL/d from 1g COD at fermenter loading with organic matter of 5.43g COD/L×d). Fermentation of sugar beet pulp gave 19% less biogas. Methane fermentation of spent hops hydrolysate yielded 121.47mL/d biogas from 1g COD (at 6.02g COD/L×d, 13% more than from spent hops). These results provide evidence that suitable enzymatic pretreatment of lignocellulosic wastes improve biogas yield from anaerobic fermentation.

Dilute H2SO4-catalyzed hydrothermal pretreatment to enhance enzymatic digestibility of Jatropha curcas fruit hull for ethanol fermentation

Dilute sulfuric acid pretreatment of the Jatropha curcas fruit hull at high temperatures (140°C to 180°C) performed in a 110-mL stainless steel reactor was investigated to enhance the enzymatic digestibility of its lignocellulosic components. Carbohydrates accounted for 43% of the dry matter of the J. curcas fruit hull biomass. The goal of the study was to optimize the pretreatment conditions (acid concentration, time, and temperature) in order to obtain the highest sugar yield after subsequent enzymatic hydrolysis. A Box-Behnken design was applied to the experimental setup in order to reduce the number of experiments. The optimal pretreatment conditions are 30-min incubations at a temperature of 178°C with a sulfuric acid concentration of 0.9% (w/v). Using these pretreatment conditions for a fruit solid loading of 9.52% followed by a 24-h enzymatic hydrolysis resulted in a liberation of 100% of all pentoses present (71% yield and 29% degradation to furfural) and 83% of the hexoses (78% yield and 5% degradation to 5-hydroxymethylfurfural). The simultaneous saccharification and fermentation experiment showed that acid-pretreated fruit hull can be used as a substrate for Saccharomyces cerevisiae to produce ethanol.

The uptake of oligogalacturonide and its effect on growth inhibition, lactate dehydrogenase activity and galactin-3 release of human cancer cells

Anti-tumour activity and membrane permeability of 1kDa oligogalacturonide (PET-pectin), prepared from citrus pectin after 24h hydrolysis, by commercial pectic enzyme produced by Aspergillus niger, on four human cell lines (HepG2, A549, Colo 205, and HEK293) and the uptake of oligogalacturonide by HEK293 cell and BALB/c mouse were investigated. PET-pectin causes a higher value of growth inhibition, lactate dehydrogenase release, and galactin-3 release in human cancer cells, as compared to the human normal HEK293 cell. There was a good correlation between growth inhibition of human cells and the uptake of rhodamine B-PET-pectin content by these cells. Additionally, almost no difference between growth inhibitions of human normal HEK293 cells cultivated with PET-pectin and pectin was found. Total pectin content in the blood of PET-pectin administrated mice increased to a maximum at 2h after oral administration, while it did not increase and change in pectin administrated mice. Our findings suggested that citrus PET-pectin can be developed as a potential dietary supplement in plant origin for cancer prevention.

Fuel Ethanol Production from Wet Oxidised Corn Stover by S. Cerevisiae

In order to find out appropriate process for ethanol production from corn stover, wet oxidation（195°C，15 minutes）and simultaneous saccharification and fermentation (SSF) was carried out to produce ethanol. The results showed that the cellulose recovery of 92.9% and the hemicellulose recovery of 74.6% were obtained after pretreatment. 86.5% of cellulose was remained in the solid cake . After 24h hydrolysis at 50°C using cellulase（Cellubrix L）,the achieved conversion of cellulose to glucose was 64.8%. Ethanol production was evaluated from dried solid cake and the hydrolysate was employed as liquid fraction . After 142 h of SSF with substrate concentration of 8% (W/V), ethanol yield of 73.1 % of the theoretical based on glucose in the raw material was obtained by S. cerevisiae(ordinary baker’ yeast) . The corresponding ethanol concentration and volumetric productivity were 17.2g/L and 0.121g/L.h respectively. The estimated total ethanol production was 257.7 kg/ton raw material by assuming consumption of both C-6 and C-5. No obvious inhibition effect occurred during SSF. These instructions give you the basic guidelines for preparing papers for WCICA/IEEE conference proceedings.

Accurate determination of the amino acid content of selected feedstuffs

The accurate determination of the amino acid content is important. In the present study, a least-squares non-linear regression model of the amino acid content determined over multiple hydrolysis times was used to accurately determine the content of amino acids in five different feedstuffs. These values were compared with 24-h hydrolysis values determined for the same feedstuffs. Overall, approximately two-thirds of the amino acids determined in this study (aspartic acid, threonine, glutamic acid, proline, glycine, alanine, leucine, tyrosine, phenylalanine and arginine) using 24-h hydrolysis were in good agreement (<3% difference). When examined across feedstuffs, the concentration of serine was underestimated by the 24-h hydrolysis method by 4.8%, while the concentrations of histidine and lysine were overestimated by 3.9% and 3.1%, respectively.

Enzymatic preparation of chitooligosaccharides by commercial lipase

The effect of a commercial lipase on chitosan degradation was investigated. When four chitosans with various degrees of deacetylation were used as substrates, the lipase showed higher optimal pH toward chitosan with higher DD (degree of deacetylation). The optimal temperature of the lipase was 55°C for all chitosans. The enzyme exhibited higher activity to chitosans which were 82.8% and 73.2% deacetylated. Kinetics experiments show that chitosans with DD of 82.8% and 73.2% which resulted in lower Km values had stronger affinity for the lipase. The chitosan hydrolysis carried out at 37°C produced larger quantity of COS (chitooligosaccharides) than that at 55°C when the reaction time was longer than 6h, and COS yield of 24h hydrolysis at 37°C was 93.8%. Products analysis results demonstrate that the enzyme produced glucosamine and chitooligosaccharides with DP (degree of polymerization) of 2–6 and above, and it acted on chitosan in both exo- and endo-hydrolytic manner.

Amino acid composition determined using multiple hydrolysis times for three goat milk formulations

The amino acid composition of goat milk formulations with varying protein and carbohydrate concentrations were determined. Proteins in goat milk infant formula, goat milk growing-up formula and goat whole milk powder were hydrolysed using multiple hydrolysis time intervals. A least-squares non-linear regression model was used to predict the free and protein bound amino acid concentrations. The amino acid composition of goat infant formula was compared with human milk reference values. There was good agreement between the multiple hydrolysis and single 24-h hydrolysis methods for approximately one-half of the amino acids. Tryptophan, aspartic acid, threonine, tyrosine, isoleucine, valine, serine and alanine contents were underestimated by 10.6, 5.6, 5.6, 4.7, 4.4, 3.7, 3.7 and 3.6%, respectively, by the single 24-h hydrolysis. The study provides accurate reference data on the amino acid composition of goat milk powders. Goat milk infant formula has amino acids in amounts similar to human milk reference values, when expressed on a per-energy basis.

Synthesis of hydroxyapatite by hydrolysis of α-Ca3(PO4)2

Conditions for hydroxyapatite (HAP) synthesis in aqueous solutions by hydrolysis of α-Ca3(PO4)2 were studied. Temperature exerts a substantial effect on the rate of α-Ca3(PO4)2 hydrolysis and also changes the morphology of the reaction products. At 40 °C, the plate-like intersecting (perpendicular to the surface of the initial particles) crystals of HAP grow. Their maximum size after the 24-h hydrolysis is 1–2 µm. Needle like HAP crystals are formed upon boiling of the suspension. The morphology observed for the HAP particles agrees well with the conclusions obtained by analysis of the kinetics of tricalcium phosphate hydrolysis.

Enzymatic Preparation of Glycerides Rich in Docosahexaenoic Acid from Thraustochytrid Single Cell Oils by Candida rugosa Lipase.

Two kinds of single cell oils from thraustochytrids including Schizochytrium limacinum SR21 and Thraustochytrium sp. KK17-3 were hydrolyzed with Candida rugosa lipase to prepare the acylglycerols rich in docosahexaenoic acid (DHA, C22:6n-3). When the lipase at 800 units/g-oil was employed for a 24-h hydrolysis of SR21 oil at 37°C, DHA content in the glycerides fraction reached at up to 67.3%, being much higher than that in the original oil (40.5%), even in the absence of surfactants. In particular, monoacylglycerol contained DHA at 85.6%. After removal of free fatty acids, the glycerides fraction was further hydrolyzed to improve its DHA content to 81.3%. On the contrary, an incapability to concentrate docosapentaenoic acid (DPA, C22:5n-6) in the glycerides suggested that the lipase strictly discriminated in fine structure of respective fatty acids. Although the addition of either Triton X-100 or deoxycholate, but not Tween 20, facilitated the rapid production of monoacylglycerol rich in DHA, its accumulation level was not significantly improved. The lipase was also effective in enriching DHA (63.0%) in glycerides fraction from KK17-3 oil (30.7%). Eicosapentaenoic acid (EPA, C20:5n-3) and arachidonic acid (AA, C20:4n-6) present in this oil were released from glycerides faster than DPA and DHA by the hydrolysis. The resistance to hydrolysis of the predominant ester bonds composed of the polyunsaturates and palmitic acid (C16:0) in the oils from SR21 and KK17-3 was in the following order, DHA>>DPA>EPA>AA>C16:0. The thraustochytrid oils are therefore suitable source for the preparation of DHA-rich glycerides, especially monoglycerides.