Cycling around Lysine Modifications.

Recent studies have revealed the existence of a plethora of previously unknown protein acyl-lysine modifications, affecting the functions of targets involved in diverse cellular processes. A recent study from the Hirschey laboratory has provided new chemical insights into the mechanisms of protein acylation.

Convenient enzymatic resolution of (R,S)-2-methylbutyric acid catalyzed by immobilized lipases.

The application of several immobilized lipases has been explored in the enantioselective esterification of (R,S)-2-methylbutyric acid, an insect pheromone precursor. With the use of Candida antarctica B, using hexane as solvent, (R)-pentyl 2-methylbutyrate was prepared in 2 h with c 40%, eep 90%, and E = 35, while Thermomyces lanuginosus leads to c 18%, eep 91%, and E = 26. The (S)-enantiomer was obtained by the use of Candida rugosa or Rhizopus oryzae (2-h reaction, c 34% and 35%, eep 75 and 49%, and E = 10 and 4, respectively). Under optimal conditions, the effect of the solvent, the molar ratio, and the nucleophile were evaluated.

Enzymatic acylation of cyanidin-3-O-glucoside with aromatic and aliphatic acid methyl ester: Structure-stability relationships of acylated derivatives.

Anthocyanins are water-soluble pigments, but they tend to be unstable in aqueous solutions. Modification of their molecular structure offers a viable approach to alter their intrinsic properties and enhance stability. Aromatic and aliphatic acid methyl esters were used as acyl donors in the enzymatic acylation of cyanidin-3-O-glucoside (C3G), and their analysis was conducted using ultraperformance liquid chromatography-mass spectrometry (UPLC-MS). The highest conversion rate achieved was 96.41 % for cyanidin-3-O-(6″-feruloyl) glucoside. Comparative evaluations of stability revealed that aromatic acyl group-conjugated C3G exhibited superior stability enhancement compared with aliphatic acyl group derivatives. The stability of aliphatic C3G decreased with increasing carbon chain length. The molecular geometries of different anthocyanins were optimized, and energy level calculations using density functional theory (DFT) identified their sites with antioxidant activities. Computational calculations aligned with the in vitro antioxidant assay results. This study provided theoretical support for stabilizing anthocyanins and broadened the application of acylated anthocyanins as food colorants and nutrient supplements.

Esterification of black bean anthocyanins with unsaturated oleic acid, and application characteristics of the product.

Esterification of anthocyanins with saturated fatty acids have been widely investigated, while that with unsaturated fatty acids is little understood. In this study, crude extract (purity âˆ¼ 35 %) of cyanidin-3-O-glucoside (C3G) from black bean seed coat was utilized as reaction substrate, and enzymatically acylated with unsaturated fatty acid (oleic acid). Optimization of various reaction parameters finally resulted in the highest acylation rate of 54.3 %. HPLC-MS/MS and NMR analyses elucidated the structure of cyanidin-3-O-glucoside-oleic acid ester (C3G-OA) to be cyanidin-3-O-(6″-octadecene)-glucoside. Introduction of oleic acid into C3G improved the lipophilicity, antioxidant ability, and antibacterial activity. Further, the color and substance stability analyses showed that the susceptibility of C3G and C3G-OA to different thermal, peroxidative, and illuminant treatments were highly pH dependent, which suggested individual application guidelines. Moreover, C3G-OA showed lower toxicity to normal cell (QSG-7701) and better inhibitory effect on the proliferation of HepG2 cells than C3G, which indicated its potential anti-tumor bioactivity.

Acylated anthocyanin inhibited the formation of heterocyclic amines in hybrid chemical model system and its underlying mechanism.

Enzymatic acylation was employed to synthesize acylated anthocyanin, and a hybrid chemical model system was used for the formation of heterocyclic amines. And the inhibition effect and underline mechanism were investigated by analyzing the variations in important precursors and intermediates. Results confirmed that cyanidin-3-(6-cinnamoyl) -glycosidase (C3(6C)G) with a purity of 98.9% was obtained. HPLC identified seven types of heterocyclic amines (IQ, MeIQx, 4, 8-DimeiqX, Norharman, Harman, PhIP, and AαC) generated in the chemical model. (C3(6C)G) showed a good concentration-dependent manner for the inhibition effect on most HCAs except for MeIQx and PhIP. It also suppressed the glucose content, showed a dose-dependent manner in creatine/creatinine inhibition, and could scavenge formaldehyde, acetaldehyde, and phenylacetaldehyde. Two potential pathways might be involved: 1. by inhibiting the content of precursors (glucose and creatinine), competing with the formation of amino acids, to suppress HCAs generation; 2 through the removal of reactive carbonyl, reducing its reaction with creatinine.

Enzymatic acylation improves the stability and bioactivity of lutein: Protective effects of acylated lutein derivatives on L-O2 cells upon H2O2-induced oxidative stress.

The instability of lutein has limited its wide application especially in the food industry. In this study, enzymatic acylation of lutein with divinyl adipate was investigated. Three new acylated lutein derivatives, lutein-3-O-adipate (compound 1), lutein-3'-O-adipate (compound 2) and lutein-di-adipate (compound 3), were identified and their stabilities and bioactivates were evaluated. Notably, compounds 1-3 showed better thermal, light stability and stronger scavenging capacity to ABTS radical cation (ABTS+) and hydroxyl radical (OH). Most importantly, these acylated lutein derivatives exhibited excellent protective effects on L-O2 cells upon hydrogen peroxide (H2O2)-induced oxidative stress. In particular, the acylated lutein derivative termed compound 3 prevented cellular oxidative stress via restraining the overproduction of reactive oxygen species (ROS), thereby increasing related antioxidant enzymes activity and inhibiting apoptosis by mitochondria pathway. Our research provides important insights into the application of acylated lutein derivatives in food, cosmetic, and pharmaceutical products.

Pharmacological Activities and Chemical Stability of Natural and Enzymatically Acylated Anthocyanins: A Comparative Review.

Anthocyanins (ANCs) are naturally occurring water-soluble pigments responsible for conferring red, blue, and purple colors to fruits, vegetables, flowers, and grains. Due to their chemical structure, they are highly susceptible to degradation by external factors, such as pH, light, temperature, and oxygen. Naturally acylated anthocyanins have proven to be more stable in response to external factors and exhibit superior biological effects as compared with their non-acylated analogues. Therefore, synthetic acylation represents a viable alternative to make the application of these compounds more suitable for use. Enzyme-mediated synthetic acylation produces derivatives that are highly similar to those obtained through the natural acylation process, with the main difference between these two pathways being the catalytic site of the enzymes involved in the synthesis; acyltransferases catalyze natural acylation, while lipases catalyze synthetic acylation. In both cases, their active sites perform the addition of carbon chains to the hydroxyl groups of anthocyanin glycosyl moieties. Currently, there is no comparative information regarding natural and enzymatically acylated anthocyanins. In this sense, the aim of this review is to compare natural and enzyme-mediated synthetic acylated anthocyanins in terms of chemical stability and pharmacological activity with a focus on inflammation and diabetes.

Enzymatic Acylation of Black Rice Anthocyanins and Evaluation of Antioxidant Capacity and Stability of Their Derivatives.

In this study, the structure of the anthocyanin fractions isolated from black rice (Oryza sativa L.) was modified by the enzyme catalysis method using caffeic acid as an acyl donor. At the same time, the effects of the acylation on the lipophilicity, antioxidant activity, and stability of black rice anthocyanins were comprehensively evaluated. The structural analyses of acylated derivatives based on ultraviolet-visible spectroscopy, Fourier-transform infrared spectroscopy, ultra-high-performance liquid chromatography-high-resolution mass spectrometry, and thermogravimetric analysis revealed that caffeic acid was efficiently grafted onto the anthocyanins of black rice through an acylated reaction, while the acylation binding site was on glucoside. When the mass ratios of anthocyanins to caffeic acid were 1:1, the A319/AVis-max value of acylated anthocyanins reached 6.37. Meanwhile, the lipophilicity of acylated derivatives was enhanced. The antioxidant capacity (DPPH and FRAP) and stability (thermal, pH, and light stability) were significantly increased. Overall, the study results provide deeper insights into controlling anthocyanin homeostasis in food processing, broadening the application of colored grain products.

Inhibitory effect of acylated anthocyanins on heterocyclic amines in grilled chicken breast patty and its mechanism.

Heterocyclic amines (HCAs) are a group of carcinogenic substances produced in protein-rich poultry meat under high-temperature. Enzymatic acylation of anthocyanins (ACNs) is a reliable way to improve their stability, and we recently found the acylated cyaniding-3-O-glucose (cyanidin-3-6-cinnamoyl-glucoside, C3(6C)G) could effective inhibit the HCAs formation, but the underline mechanism was still obscure. Thus, the present study investigated the inhibitory effect ofC3(6C)G on HCAs formation in the food system (chicken breast) and to explore the potential mechanism. The results showed that C3(6C)G with different concentrations (0.1, 0.5 and 1.0 mg/mL) could significantly inhibit lipid oxidation and decrease the total HCAs content (P<0.05) in chicken breast meat patty after roasting. The samples with 0.1 mg/mL C3(6C)G had the best inhibition effect on total HCAs, with an inhibition rate of 28%, and the inhibition rates for IQ, Harman, TRP-P-2, PhIP and AαC were 34%, 46%, 100%, 54% and 41%, respectively.

Improvement of stability and lipophilicity of pelargonidin-3-glucoside by enzymatic acylation with aliphatic dicarboxylic acid.

Dicarboxylic acids derived acylated-anthocyanins are common in nature, which can also be obtained by enzymatic acylation of anthocyanins. However, little research have focused on the properties of anthocyanins with dicarboxylic acid derivatives due to the complexity of isolation, detection, and identification. In this work, pelargonidin-3-glucoside (Pg3G) was acylated with various dicarboxylic acids. The conversion yields of acylated Pg3G were positively associated with carbon chain lengths of dicarboxylic acids. The primary acylated products were identified as pelargonidin-3-(6″-malonyl) glucoside, pelargonidin-3-(6″-succinyl) glucoside, and pelargonidin-3-(6″-glutaryl) glucoside using LC-MS and NMR. Furthermore, the three acylated Pg3G derivatives exhibited improved thermostability and enhanced lipophilicity compared with Pg3G. The improved thermostability was attributed to the influence of dicarboxylic acids substituent on the distribution of flavylium cation, quinoidal base, hemiketal, cis-chalcone, and trans-chalcone at the equilibrium condition. Overall, our research provided insights about the improved stability and lipophilicity of pelargonidin-3-glucoside following enzymatic acylation with aliphatic dicarboxylic acids.

Enzymatic acylation of raspberry anthocyanin: Evaluations on its stability and oxidative stress prevention.

Raspberry anthocyanins were isolated and purified by XAD-7HP macroporous resin and silica gel column chromatography. Anthocyanins were then acylated with methyl salicylate as catalyzed by lipase under reduced pressure, and the conversion rate was 84.26%. LC-MS and NMR were used to identify the structure, and the stability, antioxidant capacity and protective ability of the acylated anthocyanins against oxidative damage were determined. The results showed that cyanindin-3-O-glucoside (C3G) was the primary anthocyanin in raspberry, and the binding site of acylation was on the glucoside C-6, and the product was cyanidin-3-(6-salicyloyl) glucoside (C3-6(S) G). After acylation, its stability in light, heat and oxidation environments could be significantly improved, and acylated ACN showed insignificant changes in antioxidant capacities to scavenge DPPH and ABTS free radicals, as well as oxygen free radical absorptive capacity (ORAC). And it could also effectively prevent the release of ROS caused by oxidative damage and alleviate oxidative stress damage.

Enzymatic acylation of cyanidin-3-O-glucoside in raspberry anthocyanins for intelligent packaging: Improvement of stability, lipophilicity and functional properties.

Anthocyanins (ACNs) as one category of water-soluble flavonoid pigments are increasingly employed in pH sensing indicator applications for monitoring food freshness. Nevertheless, considering that anthocyanins are sensitive to environmental factors, their practical applications in food industries are still rather limited. In order to improve the stability of anthocyanins and capitalize upon their application in pH-color responsive intelligent packaging, this study aims to graft octanoic aid onto raspberry anthocyanins catalyzed by immobilized Candida antarctica lipase B (Novozymes 435). Structural analyses based on Fourier transform infrared spectroscopy (FTIR), UV-Vis, liquid chromatography-mass spectrometry (LC-MS), and nuclear magnetic resonance (NMR) revealed that octanoic acid was regioselective grafted onto the 6-OH position of its glucoside. The acylation efficiency of C3G by octanoic acid up to 47.1%. The octanoic acid moiety was found to improve lipophilicity, antioxidant activity and stability of C3G. In addition, acylated derivative also maintained the pH-color response characteristics of nature ACNs and exhibited excellent NH3 response properties. These results indicated that acylated anthocyanins exhibit potential application as intelligent packaging indicator for monitoring food freshness.

Combining the mechanical ball milling of the carbohydrate and the use of low solvent reaction media for the synthesis of fructose fatty acid esters by immobilized lipases.

Synthesis of carbohydrate fatty acid esters catalyzed by immobilized lipases is a pathway to obtain specific isomers from renewable feedstock, compared to unselective chemical esterification. While the use of low-solvent reaction media (≤ 10 %) offers advantages, the interactive effects of these media with biocatalysts and substrates should be modulated towards high catalytic efficiency and substrate availability. Among the investigated co-solvents, tert-butanol and DMSO in a mixture of lauric acid substrate/co-solvent (90/10; v/v) resulted in high bioconversion yields using either Novozym® 435 or Lipozyme® RM IM, as biocatalysts. Increased hydrophobicity of the Novozym® 435 immobilization support favored bioconversion, while polar substrate surface area enlargement by ball-milling improved productivity through enhancement of fructose availability. A compromise between bioconversion yield (19.7 %) and productivity (9.45 µmol/L min) was obtained in the reactions catalyzed by Novozym® 435 using ball-milled fructose at a concentration of 0.2 mol/L. Combining mechanical ball-milling of the substrates with low-solvent reaction media is expected to enhance and expand enzymatic synthesis of carbohydrate fatty acid esters.

Enzymatic acylation of rutin with benzoic acid ester and lipophilic, antiradical, and antiproliferative properties of the acylated derivatives.

Rutin (3',4',5,7-tetrahydroxy-flavone-3-rutinoside) was enzymatically acylated with benzoic acid and its esters (methyl benzoate and vinyl benzoate) using Thermomyces lanuginosus lipase (Lipozyme TLIM). The acylation reaction was optimized by varying the reaction medium, reaction temperature, acyl donor, substrate molar ratio, and reaction time. The highest conversion yield (76%) was obtained in tert-amyl alcohol (60 °C, 72 hr) using vinyl benzoate (molar ratio of 1:10) as acyl donor. The acylation occurred at the 2'''-OH and 4'''-OH of the rhamnose unit and the 2''-OH position of the glucose moieties. Three novel rutin acylated derivatives (compounds 1-3) were purified and characterized by HR-MS and 1D and 2D NMR spectroscopy. We found that acylation significantly improved lipophilicity, capacity to inhibit lipid peroxidation, anticancer capacity and substantially maintained the antioxidant activity of rutin. This research provides important insights in the acylation of flavonoids with different glycosyl moieties. PRACTICAL APPLICATION: In this study, three novel rutin derivatives were successfully synthesized and the highest conversion yield (76%) was obtained by reacting the rutin and vinyl benzoate at molar ratio of 1:10 in tert-amyl alcohol for 72 hr at 60 °C. Introducing a benzoic acid substituent into rutin molecule significantly improved their lipophilicity and inhibition of lipid peroxidation in lipophilic system. Furthermore, this study demonstrated that acylation significantly improved anticancer capacity and substantially maintained the antioxidant activity.

Efficient Enzymatic Preparation of Flavor Esters in Water.

A straightforward biocatalytic method for the enzymatic preparation of different flavor esters starting from primary alcohols (e.g., isoamyl, n-hexyl, geranyl, cinnamyl, 2-phenethyl, and benzyl alcohols) and naturally available ethyl esters (e.g., formate, acetate, propionate, and butyrate) was developed. The biotransformations are catalyzed by an acyltransferase from Mycobacterium smegmatis (MsAcT) and proceeded with excellent yields (80-97%) and short reaction times (30-120 min), even when high substrate concentrations (up to 0.5 M) were used. This enzymatic strategy represents an efficient alternative to the application of lipases in organic solvents and a significant improvement compared with already known methods in terms of reduced use of organic solvents, paving the way to sustainable and efficient preparation of natural flavoring agents.

Enzymatic acylation of flavone isolated from extractive of bamboo leaves with oleic acid and antioxidant activity of acylated product.

This present study aims to establish a relatively cheap and effective method for preparation of ester of flavone isolated from bamboo-leaf extracts. According to this method, the solubility of bamboo leaf extract in a lipid medium could be improved, which would expand the utilization of bamboo leaf extract. Acylation of flavone with oleic acid was efficient in a system containing dried tert-amyl-alcohol and molecular sieves. The bioconversion yield ranged from 24 to 63% in different conditions. the type of fatty acids, the amount of the lipase, water content and temperature of the system were investigated in detail to influence on the performance of the acylation reaction. High purity of ester (>95%) was obtained by a two-step solvent purification procedure and chromatographic separation. Followed by the 1H and 13C nuclear magnetic resonance spectroscopy analysis, single product of 6''-O-oleate isoorientin ester was detected. Introducing an acyl group into isoorientin significantly improved its lipophilicity yet slightly reduced their antioxidant activity, while its antioxidant activity still significantly better than VE.

Enzymatic acylation of blackcurrant (Ribes nigrum) anthocyanins and evaluation of lipophilic properties and antioxidant capacity of derivatives.

Anthocyanin-rich fractions isolated from blackcurrant (Ribes nigrum L.) including delphinidin-3-O-glucoside, delphinidin-3-O-rutinoside, cyanidin-3-O-glucoside and cyanidin-3-O-rutinoside were enzymatically acylated with lauric acid. All the four anthocyanins were successfully monoacylated, and their relative proportions did not affect the conversion yield. The acylation occurred at the 6″-OH position of the glucosides and at the rhamnose 4‴-OH of the rutinosides. The rutinoside moieties of the anthocyanins were successfully acylated for the first time, and the corresponding acylation sites were verified by NMR analysis. The acylation enhanced the lipophilicity. The hydrophilic anthocyanin rutinosides were more lipophilic after acylation. Introducing lauric acid into the anthocyanins significantly improved the thermostability and capacity to inhibit lipid peroxidation and maintained UV-vis absorbance and antioxidant activity. This research provides important insights into acylation of mixed anthocyanins with different glycosyl moieties.

Enzymatic Acylation of Anthocyanin Isolated from Black Rice with Methyl Aromatic Acid Ester as Donor: Stability of the Acylated Derivatives.

The enzymatic acylation of anthocyanin from black rice with aromatic acid methyl esters as acyl donors and Candida antarctica lipase B was carried out under reduced pressure. The highest conversion of 91% was obtained with benzoic acid methyl ester as acyl donor; cyanidin 3-(6″-benzoyl)-glucoside, cyanidin 3-(6″-salicyloyl)-glucoside, and cyanidin 3-(6″-cinnamoyl)-glucoside were successfully synthesized. This is the first report on the enzymatic acylation of anthocyanin from black rice with methyl aromatic esters as acyl donors and lipase as biocatalyst. Furthermore, the acylation with aromatic carboxylic acids enhanced both the thermostability and light resistivity of anthocyanin. In particular, cyanidin 3-(6″-cinnamoyl)-glucoside was the most stable among the three acylated anthocyanins synthesized.

Efficient Regioselective Synthesis of the Crotonyl Polydatin Prodrug by Thermomyces lanuginosus Lipase: a Kinetics Study in Eco-friendly 2-Methyltetrahydrofuran.

Bio-based solvents have recently been discussed as sustainable green and promising alternatives to conventional organic media for enzymatic processes. In this paper, highly regioselective synthesis of the 6″-O-crotonyl-polydatin catalyzed by Thermomyces lanuginosus lipase (TLL) in biomass-derived 2-methyltetrahydrofuran (2-MeTHF) was successfully performed for the first time. The results indicated that TLL lipase displayed significantly improved catalytic performance in 2-MeTHF than in other traditional solvents. Under the optimal conditions, the initial reaction rate, 6″-regioselectivity, and maximum substrate conversion were as high as 12.38 mM h(-1), 100 %, and 100 %, respectively. Moreover, further investigations on the operational stability, kinetic parameters like V max, K m, V max/K m, and E a revealed that 2-MeTHF exhibited excellent biocompatibility and rendered the greener process of the enzymatic acylation.

Synthesis and emulsifying properties of carbohydrate fatty acid esters produced from Agave tequilana fructans by enzymatic acylation.

Carbohydrate fatty acid esters are non-ionic surfactants with a broad spectrum of applications. These molecules are generally synthesized using short carbohydrates or linear fructans; however in this research carbohydrate fatty acid esters were produced for the first time with branched fructans from Agave tequilana. Using immobilized lipases we successfully acylated A. tequilana fructans with vinyl laurate, obtaining products with different degrees of polymerization (DP). Lipozyme 435 was the most efficient lipase to catalyze the transesterification reaction. HPLC and ESI-MS analysis proved the presence of a mixture of acylated products as a result of the chemical complexity of fructans in the A. tequilana. The ESI-MS spectra showed a molecular mass shift between 183 and 366g/mol for fructooligosaccharides with a DP lower than 6, which indicated the presence of Agave fructans that had been mono- and diacylated with lauric acid. The carbohydrate fatty acid esters (CFAE) obtained showed good emulsifying properties in W/O emulsions.