脂肪酸光脱羧酶及其在有机合成中应用的研究进展

生物基脂肪酸是极具应用潜力的可再生资源。以绿色、低碳的方式对生物基脂肪酸进行转化合成高价值的化学品和燃料可满足现代社会可持续发展的迫切需要,也符合绿色生物制造的需求。生物催化因反应条件温和、选择性高等特点受到众多学者的广泛关注,其中,近年来新发现的由可见光驱动的脂肪酸光脱羧酶（fatty acid photodecarboxyase, CvFAP）可催化不同链长的脂肪酸进行脱羧反应用来合成燃料和化学品,反应过程具有极高原子经济性,在绿色催化工艺及能源领域具有广阔的应用前景。本综述旨在对脂肪酸光脱羧酶（CvFAP）的最新研究进展进行总结分析,并对脂肪酸光脱羧酶的发展趋势进行展望,以期为脂肪酸光脱羧酶（CvFAP）在绿色合成领域的进一步开发利用提供参考。     

CYP505E3: A Novel Self‐Sufficient ω‐7 In‐Chain Hydroxylase

The self‐sufficient cytochrome P450 monooxygenase CYP505E3 from Aspergillus terreus catalyzes the regioselective in‐chain hydroxylation of alkanes, fatty alcohols, and fatty acids at the ω‐7 position. It is the first reported P450 to give regioselective in‐chain ω‐7 hydroxylation of C10–C16 n‐alkanes, thereby enabling the one step biocatalytic synthesis of rare alcohols such as 5‐dodecanol and 7‐tetradecanol. It shows more than 70 % regioselectivity for the eighth carbon from one methyl terminus, and displays remarkably high activity towards decane (TTN≈8000) and dodecane (TTN≈2000). CYP505E3 can be used to synthesize the high‐value flavour compound δ‐dodecalactone via two routes: 1) conversion of dodecanoic acid into 5‐hydroxydodecanoic acid (24 % regioselectivity), which at low pH lactonises to δ‐dodecalactone, and 2) conversion of 1‐dodecanol into 1,5‐dodecanediol (55 % regioselectivity), which can be converted into δ‐dodecalactone by horse liver alcohol dehydrogenase.     

Stabilizing catalytic pathways via redundancy: selective reduction of microalgae oil to alkanes

A new route to convert crude microalgae oils using ZrO(2)-promoted Ni catalysts into diesel-range alkanes in a cascade reaction is presented. Ni nanoparticles catalyze the selective cleavage of the C-O of fatty acid esters, leading to the hydrogenolysis of triglycerides. Hydrogenation of the resulting fatty acids to aldehydes (rate-determining step) is uniquely catalyzed via two parallel pathways, one via aldehyde formation on metallic Ni and the second via a synergistic action by Ni and ZrO(2) through adsorbing the carboxylic groups at the oxygen vacancies of ZrO(2) to form carboxylates and subsequently abstracting the α-hydrogen atom to produce ketene, which is in turn hydrogenated to aldehydes and decarbonylated on Ni nanoparticles.     

Sulfide Catalysts for Production of Motor Fuels from Fatty Acid Triglycerides

Patents dealing with the production of motor fuel components by hydrodeoxygenation of renewable raw materials based on fatty acid triglycerides are analyzed. Various methods of using sulfide catalysts in hydrodeoxygenation of fatty acid triglycerides and of their mixtures with petroleum fractions are described. The ways to overcome problems that arise in hydrodeoxygenation, based on using sulfide catalysts differing in the active component and support composition, are considered. For example, the use of supported MoS₂ catalysts free of Co and Ni ensures the conversion of fatty acid triglycerides along the “direct hydrodeoxygenation” pathway to avoid the formation of carbon oxides and related process problems. The use of sulfide catalysts on zeolite-containing supports allows synthesis of products with improved low-temperature properties due to isomerization (or mild hydrocracking) of С₁₅–С₁₈ alkanes formed by hydrodeoxygenation of fatty acid triglycerides.

     

Quantitative analysis of fatty-acid-based biofuels produced by wild-type and genetically engineered cyanobacteria by gas chromatography-mass spectrometry

Simple and rapid quantitative determination of fatty-acid-based biofuels is greatly important for the study of genetic engineering progress for biofuels production by microalgae. Ideal biofuels produced from biological systems should be chemically similar to petroleum, like fatty-acid-based molecules including free fatty acids, fatty acid methyl esters, fatty acid ethyl esters, fatty alcohols and fatty alkanes. This study founded a gas chromatography-mass spectrometry (GC-MS) method for simultaneous quantification of seven free fatty acids, nine fatty acid methyl esters, five fatty acid ethyl esters, five fatty alcohols and three fatty alkanes produced by wild-type Synechocystis PCC 6803 and its genetically engineered strain. Data obtained from GC-MS analyses were quantified using internal standard peak area comparisons. The linearity, limit of detection (LOD) and precision (RSD) of the method were evaluated. The results demonstrated that fatty-acid-based biofuels can be directly determined by GC-MS without derivation. Therefore, rapid and reliable quantitative analysis of fatty-acid-based biofuels produced by wild-type and genetically engineered cyanobacteria can be achieved using the GC-MS method founded in this work.     

Phase and chemical equilibria in the transesterification reaction of vegetable oils with supercritical lower alcohols

Calculations of thermodynamic data are performed for fatty acid triglycerides, free fatty acids, and fatty acid methyl esters, participants of the transesterification reaction of vegetable oils that occurs in methanol. Using the obtained thermodynamic parameters, the phase diagrams for the reaction mixture are constructed, and the chemical equilibria of the esterification reaction of free fatty acids and the transesterification reaction of fatty acid triglycerides attained upon treatment with supercritical methanol are determined. Relying on our analysis of the obtained equilibria for the esterification reaction of fatty acids and the transesterification reaction of triglycerides attained upon treatment with lower alcohols, we select the optimum conditions for performing the reaction in practice.     

Can gas chromatography combustion isotope ratio mass spectrometry be used to quantify organic compound abundance?

Quantifying the concentrations of organics such as phospholipid fatty acids (PLFAs) and n‐alkanes and measuring their corresponding ¹³ C/¹² C isotope ratios often involves two separate analyses; (1) quantification by gas chromatography flame ionisation detection (GC‐FID) or gas chromatography/mass spectrometry (GC/MS), and (2) ¹³ C‐isotope abundance analysis by gas chromatography/combustion/isotope ratio mass spectrometry (GC‐C‐IRMS). This requirement for two separate analyses has obvious disadvantages in terms of cost and time. However, there is a history of using the data output of isotope ratio mass spectrometers to quantify various components; including the N and C concentrations of solid materials and CO₂ concentrations in gaseous samples. Here we explore the possibility of quantifying n‐alkanes extracted from sheeps' faeces and fatty acid methyl esters (FAMEs) derivatised from PLFAs extracted from grassland soil, using GC‐C‐IRMS. The results were compared with those from GC‐FID analysis of the same extracts. For GC‐C‐IRMS the combined area of the masses for all the ions (m/z 44, 45 and 46) was collected, referred to as 'area all', while for the GC‐FID analysis the peak area data were collected. Following normalisation to a common value for added internal standards, the GC‐C‐IRMS 'area all' values and the GC‐FID peak area data were directly compared. Strong linear relationships were found for both n‐alkanes and FAMEs. For the n‐alkanes the relationships were 1:1 while, for the FAMEs, GC‐C‐IRMS overestimated the areas relative to the GC‐FID results. However, with suitable reference material 1:1 relationships were established. The output of a GC‐C‐IRMS system can form the basis for the quantification of certain organics including FAMEs and n‐alkanes. Copyright © 2011 John Wiley & Sons, Ltd.     

Development of saw palmetto (Serenoa repens) fruit and extract standard reference materials

As part of a collaboration with the National Institutes of Health’s Office of Dietary Supplements and the Food and Drug Administration’s Center for Drug Evaluation and Research, the National Institute of Standards and Technology has developed two standard reference materials (SRMs) representing different forms of saw palmetto (Serenoa repens), SRM 3250 Serenoa repens fruit and SRM 3251 Serenoa repens extract. Both of these SRMs have been characterized for their fatty acid and phytosterol content. The fatty acid concentration values are based on results from gas chromatography with flame ionization detection (GC-FID) and mass spectrometry (GC/MS) analysis while the sterol concentration values are based on results from GC-FID and liquid chromatography with mass spectrometry analysis. In addition, SRM 3250 has been characterized for lead content, and SRM 3251 has been characterized for the content of β-carotene and tocopherols. SRM 3250 (fruit) has certified concentration values for three phytosterols, 14 fatty acids as triglycerides, and lead along with reference concentration values for four fatty acids as triglycerides and 16 free fatty acids. SRM 3251 (extract) has certified concentration values for three phytosterols, 17 fatty acids as triglycerides, β-carotene, and γ-tocopherol along with reference concentration values for three fatty acids as triglycerides, 17 fatty acids as free fatty acids, β-carotene isomers, and δ-tocopherol and information values for two phytosterols. These SRMs will complement other reference materials currently available with concentrations for similar analytes and are part of a series of SRMs being developed for dietary supplements. Contribution of the US Government; not subject to copyright     

Analysis of triglycerides in food items by desorption electrospray ionization mass spectrometry

The triglyceride composition and oxidation behavior of edible oil and margarine samples were analyzed by desorption electrospray ionization mass spectrometry (DESI‐MS). For the characterization of the lipids, the chain length and the degree of unsaturation of the fatty acids were determined. The measurements were carried out in positive ion mode; the triglycerides were detected as alkali metal or ammonium adducts. The DESI solvent was water/methanol 1:1 (v/v); measurements were carried out both with and without the addition, as an ionizing agent, of ammonium acetate that enhances the signal intensity of the ammonium adduct ions. The spectra were interpreted for both cases and intensities were compared. Triglyceride monomers and dimers were observed in the spectra. Tandem mass spectrometric (MS/MS) measurements were carried out to determine the structure of the triglycerides. It was demonstrated that the terminal fatty acids in the sn1‐ or sn3‐position are more likely to be cleaved than the internal fatty acid (sn2‐position). Characteristic triglyceride patterns were obtained using a simple and rapid sample preparation protocol comprising the simple deposition of samples onto a glass carrier surface. The triglyceride data was analyzed by principal component analysis (PCA). The different edible oils were clearly separated and the hydrogenated derivatives were identified by their triglyceride spectra. The oxidation of the oil samples was observed and the oxidation products were detected and identified. This method provides a fast and simple technique for the detection and analysis of triglycerides in oil‐ or fat‐containing samples ranging from food items to tissue samples. The potential application areas include nutritional studies, the food industry and cosmetics. Copyright © 2010 John Wiley & Sons, Ltd.     

Light-Driven Enzymatic Decarboxylation of Dicarboxylic Acids

Photodecarboxylase from Chlorella variabillis (CvFAP) is one of the three known light-activated enzymes that catalyzes the decarboxylation of fatty acids into the corresponding C1-shortened alkanes. Although the substrate scope of CvFAP has been altered by protein engineering and decoy molecules, it is still limited to mono-fatty acids. Our studies demonstrate for the first time that long chain dicarboxylic acids can be converted by CvFAP. Notably, the conversion of dicarboxylic acids to alkanes still represents a chemically very challenging reaction. Herein, the light-driven enzymatic decarboxylation of dicarboxylic acids to the corresponding (C2-shortened) alkanes using CvFAP is described. A series of dicarboxylic acids is decarboxylated into alkanes in good yields by means of this approach, even for the preparative scales. Reaction pathway studies show that mono-fatty acids are formed as the intermediate products before the final release of C2-shortened alkanes. In addition, the thermostability, storage stability, and recyclability of CvFAP for decarboxylation of dicarboxylic acids are well evaluated. These results represent an advancement over the current state-of-the-art.     

Analysis of tissue free fatty acids isolated by aminopropyl bonded-phase columns

Gas chromatographic analysis revealed that polyunsaturated fatty acids such as arachidonic acid and total tissue free fatty acids isolated from an aminopropyl bonded-phase column yield a two- to three-fold higher recovery of arachidonic acid as compared to those isolated from thin-layer chromatographic plates. This method was further improved by packing the aminopropyl bonded phase in glass columns, since the glass column significantly eliminated the other contaminants (from polypropylene columns) coeluting with fatty acids in both a neutral lipid thin-layer chromatographic system and on a 5% DEGS-PS column of gas chromatographic analysis. In aminopropyl bonded-phase columns, the standard triglycerides and phospholipids were completely separated from free fatty acids as judged by gas chromatographic analysis. These results warrant the use of an aminopropyl bonded-phase column for the isolation of free fatty acids to obtain better recovery of polyunsaturated fatty acids.     

Neutral Lipids of Ruta graveolens L. Grown in vivo and in vitro

The composition of neutral lipids (NL) and fatty acids of acyl-containing components in differentiated tissues ofRuta graveolensL. (Rutaceae) grown in vivo and in vitro is studied. The NL of in vitro cultures differ from NL of the intact plant by a larger content of esters of fatty acids with alcohols, fewer high-molecular-weight alkanes, alkylbenzenes, free alkanols, and linolenic acid     

Minor gamma-lactones from Trichilia catigua (Meliaceae) and its precursors by GC-MS

NMR and GC-MS analysis of fractions of the CHCl3 extract of Trichilia catigua bark led to the identification of a mixture of three omega-phenyl alkanes, three omega-phenyl alkanoic acids, five omega-phenyl-gamma-lactones, two alkyl-gamma-lactones, one alkenyl-gamma-lactone and a mixture of fatty acids ranging from C-14 to C-26. Beta-sitosterol, stigmasterol and campesterol as free alcohols were also identified.     

Organic compounds in precipitation

Summary Precipitation samples collected in Hannover (Germany) mainly in 1989 (and in part also in 1988 and 1990) were analyzed for n-alkanes, fatty acids, aldehydes, phenols and polycyclic aromatic hydrocarbons. The analytical methods employed were: GC/FID for alkanes, GC/MS for fatty acids and phenols, HPLC for aldehydes and PAHs.     

The impact of insecticides containing deltamethrin and cyfluthrin on the composition of surface compounds in the larvae,females and males of Tenebrio molitor

This paper presents the effect of insecticides on the composition of the surface compounds of one of the most harmful insects, Tenebrio molitor, by analysis using GC–MS. As a result of the use of insecticides, the composition of the chemical compounds on the surface of insects changes, depending on the insecticides used. The most numerous groups of the marked compounds were fatty acids, alkanes, esters and sterols. The content of the identified compounds in the larvae increased at both 24 and 48 h after the application of insecticides, in comparison with the control samples. The content of identified compounds in the samples taken from the females increased 24, 48 and 72 h after the application of insecticides in comparison with the control samples. By contrast, in samples prepared from males, the content of identified compounds decreased 24 h after the application of insecticides, compared with the control samples. The highest content of chemical compounds was for fatty acids and alkanes after the use of insecticides. The content of fatty acids after the application of the insecticide with deltamethrin was 62.1 ± 3.3–466.9 ± 5.9 μg/g, and after the application of the insecticide with cyfluthrin was 49.9 ± 1.9–458.3 ± 4.2 μg/g. However, the content of alkanes after the use of deltamethrin was 115.6 ± 4.2–4672.0 ± 32.1 μg/g, and after the use of cyfluthrin was 189.4 ± 3.8–3975.0 ± 10.2 μg/g.     

Calibration of thin-layer chromatography with flame ionization detection for the analysis of natural lipid samples

Thin-layer chromatography coupled with flame ionization detection has been used to develop a method to quantitate fish lipids. Quantitation of fractionated lipid classes is usually accomplished through calibration with standards. Our work shows that various standards within a class, such as triglycerides or free fatty acids give substantially different responses to the detector. A method has been developed in which a composite sample of salmon lipid is prepared with an internal standard. This technique eliminates the variable detector response observed for compounds within a class and provides more accurate quantitation of oils from which the calibration samples were prepared. The lipid classes quantitated were triglycerides, free fatty acids, phosphatidylethanolamine and phosphatidylcholine.     

Calculating the thermodynamic characteristics of the stepwise transesterification of simple triglycerides

Thermodynamic data for mono- and diglycerides of palmitic, oleic, and linoleic fatty acids participating in the stepwise transesterification reaction of the corresponding simple triglycerides in methanol are calculated. The obtained thermodynamic parameters allow us to calculate the chemical equilibrium and the equilibrium composition of the products of the stepwise transesterification reaction of fatty acid triglycerides with supercritical methanol.     

Synthetic Polyester from Algae Oil

Current efforts to technically use microalgae focus on the generation of fuels with a molecular structure identical to crude oil based products. Here we suggest a different approach for the utilization of algae by translating the unique molecular structures of algae oil fatty acids into higher value chemical intermediates and materials. A crude extract from a microalga, the diatom Phaeodactylum tricornutum, was obtained as a multicomponent mixture containing amongst others unsaturated fatty acid (16:1, 18:1, and 20:5) phosphocholine triglycerides. Exposure of this crude algae oil to CO and methanol with the known catalyst precursor [{1,2‐(tBu₂PCH₂)₂C₆H₄}Pd(OTf)](OTf) resulted in isomerization/methoxycarbonylation of the unsaturated fatty acids into a mixture of linear 1,17‐ and 1,19‐diesters in high purity (>99 %). Polycondensation with a mixture of the corresponding diols yielded a novel mixed polyester‐17/19.17/19 with an advantageously high melting and crystallization temperature.     

Production of biodiesel by immobilized Candida sp. lipase at high water content

A new process for enzymatic synthesis of biodiesel at high water content (10–20%) with 96% conversion by lipase from Candida sp. 99–125 was studied. The lipase, a no-position-specific lipase, was immobilized by a cheap cotton membrane and the membrane-immobilized lipase could be used at least six times with high conversion. The immobilized lipase could be used for different oil conversion and preferred unsaturated fatty acids such as oleic acid to staturated fatty acids such as palmitic acid. The changes in concentration of fatty acids, diglycerides, and methyl esters in the reaction were studied and a mechanism of synthesis of biodiesel was suggested: the triglycerides are first enzymatically hydrolyzed into fatty acids, and then these fatty acids are further converted into methyl esters.     

Capillary GC of triglycerides in fats and oils using a high temperature phenylmethylsilicone stationary phase,part I

The analysis of triglycerides on FSOT columns coated with a phenylmethylsilicone gum opens up new perspectives in fat and oil characterization. On phenylmethylsilicone, besides a “carbon number” or CN separation, the triglycerides are also separated according to the different combinations of saturated and unsaturated fatty acids in the triglycerides.