Search

Brewer's spent grain (BSG), the most abundant by-product of the beer industry, can be considered a sustainable source of ferulic acid (4-hydroxy-3-methoxycinnamic acid) (FA). FA is esterified to the arabinoxylan in the BSG. FA exhibits a variety of biological activities and is a target for valorization of BSG. We present a novel enzyme-assisted approach that utilizes the synergistic action of a specific bacterial endo-1,4-β-xylanase and a fungal ferulic acid esterase activity to selectively release high yields of FA from BSG. The study compares the efficacy of three different fungal feruloyl esterases (E.C. 3.1.1.73) of carbohydrate esterase family 1 (CE1), Hi Fae, Ao Fae1, and An Fae1, acting separately or in combination with two different endo-1,4-β-xylanases (EC 3.2.1.8), a bacterial GH30 enzyme from Dickeya chrysanthemi , and Shearzyme®, a commercial GH10 enzyme from Aspergillus aculeatus , respectively. On an equimolar dosage basis, the An Fae1-GH30 was the most efficient combination in releasing FA from BSG. This combination showed significant synergy in a dose-dependent manner, enabling up to 85% FA recovery from BSG within 30 min. of treatment. The results demonstrate a novel enzymatic approach for valorization of BSG as FA is considered a high-value natural compound for use in e.g. anti-aging skin care products. [Display omitted] • Ferulic acid was effectively enzymatically released from brewers' spent grain. • Combination of An Fae1 and a GH30 xylanase liberated ferulic acid by synergism. • 85% yield of ferulic acid was liberated in 1 h at optimum reaction conditions. • Kinetics of enzymatic ferulic acid release from arabinoxylan by FAEs was studied. [ABSTRACT FROM AUTHOR]

Purpose of the review: Ferulic acid (FA), which occurs naturally as the feruloylated sugar ester in grains, fruits, and vegetables, is critical for combating oxidative stress and alleviating neurodegenerative diseases resulting from free radical-generated protein aggregates in brain cells. However, FA cannot be absorbed in conjugated form. Therefore, strategies to improve the bioavailability of FA are gaining more importance. Ferulic acid esterases (FAE) of the gut microbiota are critical enzymes that facilitate FA release from feruloylated sugar ester conjugates and influence systemic health. This review provides insight into a nutrition-based approach to preventing neurodegenerative disorders such as Alzheimer's and Parkinson's by altering the diversity of FAE-producing gut microbiota. Recent findings: The human gut is a niche for a highly dense microbial population. Nutrient components and the quality of food shape the gut microbiota. Microbiota–diet–host interaction primarily involves an array of enzymes that hydrolyse complex polysaccharides and release covalently attached moieties, thereby increasing their bio-accessibility. Moreover, genes encoding polysaccharide degrading enzymes are substrate inducible, giving selective microorganisms a competitive advantage in scavenging nutrients. Summary: Nutraceutical therapy using specific food components holds promise as a prophylactic agent and as an adjunctive treatment strategy in neurotherapeutics, as it results in upregulation of polysaccharide utilisation loci containing fae genes in the gut microbiota, thereby increasing the release of FA and other antioxidant molecules and combat neurodegenerative processes. [ABSTRACT FROM AUTHOR]

Corn stover (CS) is an abundant lignocellulosic by-product of the grain industry. Ferulic acid esterase producing (FAE+)-lactobacilli can potentially improve ensiled forages' nutritive value through the hydrolysis of ferulic acid ester bonds present in cell walls during the fermentation process, but this has not been addressed in CS silage. In this study, we characterized 8 FAE+ lactobacilli regarding their FAE activity and inoculant aptitude: Lactobacillus (L.) johnsonii (CRL2237, CRL2238, CRL2240), L. plantarum (ETC182, CRL046, CRL2241), L. fermentum CRL1446 and L. brevis CRL2239. Next, 25% dry matter (DM) CS mini silos were prepared and either not inoculated (UN) or inoculated with each strain (105 CFU g fresh matter−1). Compared to UN, DM loss was significantly reduced in CRL046 and CRL2239, and organic matter increased in CRL2241-inoculated silages. Although the rest of the digestibility measures were not improved, in situ acid detergent fiber degradability (ADFD) was increased by the CRL2238 strain when compared to UN. Results in inoculated silages were not correlated with FAE activity quantification or growth/acidification studies in a CS-derived culture broth. This study demonstrates the potential of several FAE+ lactobacilli strains as CS inoculants and encourages further research. [ABSTRACT FROM AUTHOR]

Corn stover (CS) is an abundant lignocellulosic by-product of the grain industry. Ferulic acid esterase producing (FAE+)-lactobacilli can potentially improve ensiled forages’ nutritive value through the hydrolysis of ferulic acid ester bonds present in cell walls during the fermentation process, but this has not been addressed in CS silage. In this study, we characterized 8 FAE+ lactobacilli regarding their FAE activity and inoculant aptitude: Lactobacillus (L.) johnsonii (CRL2237, CRL2238, CRL2240), L. plantarum (ETC182, CRL046, CRL2241), L. fermentum CRL1446 and L. brevis CRL2239. Next, 25% dry matter (DM) CS mini silos were prepared and either not inoculated (UN) or inoculated with each strain (105 CFU g fresh matter−1). Compared to UN, DM loss was significantly reduced in CRL046 and CRL2239, and organic matter increased in CRL2241-inoculated silages. Although the rest of the digestibility measures were not improved, in situ acid detergent fiber degradability (ADFD) was increased by the CRL2238 strain when compared to UN. Results in inoculated silages were not correlated with FAE activity quantification or growth/acidification studies in a CS-derived culture broth. This study demonstrates the potential of several FAE+ lactobacilli strains as CS inoculants and encourages further research.

Feruloyl esterases (FAEs) have an important role in the enzymatic conversion of lignocellulosic biomass by decoupling plant cell wall polysaccharides and lignin. Moreover, FAEs release anti-oxidative hydroxycinnamic acids (HCAs) from biomass. As a plethora of FAE candidates were found in fungal genomes, FAE classification related to substrate specificity is an indispensability for selection of most suitable candidates. Hence, linking distinct substrate specificities to a FAE classification, such as the recently classified FAE subfamilies (SF), is a promising approach to improve the application of these enzymes for a variety of industrial applications. In total, 14 FAEs that are classified members of SF1, 5, 6, 7, 9, and 13 were tested in this research. All FAEs were investigated for their activity toward a variety of substrates: synthetic model substrates, plant cell wall-derived substrates, including lignin, and natural substrates. Released HCAs were determined using reverse phase-ultra high performance liquid chromatography coupled to UV detection and mass spectrometry. Based on this study, FAEs of SF5 and SF7 showed the highest release of FA, pCA, and diFAs over the range of substrates, while FAEs of SF6 were comparable but less pronounced for diFAs release. These results suggest that SF5 and SF7 FAEs are promising enzymes for biorefinery applications, like the production of biofuels, where a complete degradation of the plant cell wall is desired. In contrast, SF6 FAEs might be of interest for industrial applications that require a high release of only FA and pCA, which are needed as precursors for the production of biochemicals. In contrast, FAEs of SF1, 9 and 13 showed an overall low release of HCAs from plant cell wall-derived and natural substrates. The obtained results substantiate the previous SF classification as a useful tool to predict the substrate specificity of FAEs, which eases the selection of FAE candidates for industrial applications.

In the present study, the hydrolysis and acyltransferase activities of bifunctional feruloyl esterases DLFae4 were enhanced by a combination of multiple mutation strategies based on rationally predicting the potential beneficial mutation sites. Results showed that V149D&I172G was the strongest double mutant (589.67 U/mg), and the triple mutant N173G&N169G&I148G exhibited 3.60 and 5.57 times higher hydrolytic activity and catalytic efficiency towards MFA, respectively. Furthermore, the triple mutant releases 16.16 mg/g ferulic acid (FA) from de-starched wheat bran (DSWB) in synergy with xylanase, which was increased by 9.34 times than that released by DLFae4 alone, and was the highest reported release amount. The double mutant V251Y&A341G with weakened hydrolytic activity but exhibited 4.30 times higher acyltransferase activity than that of DLFae4, and the corresponding acyl transfer efficiency improved by 11.37 times, representing the highest acyltransferase activity to date. The results from the current investigation demonstrate that combining multiple rational design methods is a robust pipeline for improving enzyme activities and provides new insights for enzyme modification. [Display omitted] [ABSTRACT FROM AUTHOR]

Genes encoding six feruloyl esterases (FAEs; lbff 0997, lbff 0272, lbff 1432, lbff 1695, lbff 1849, lbff 0153) from Lactobacillus fermentum JN248 were cloned, overexpressed and characterised. Maximum enzyme activity was observed at 35 °C for recombinant FAEs LFFae0997, LFFae0272 and LFFae0153, at 30 °C for LFFae1695, and at 40 °C for LFFae1432and LFFae1849. For five of the enzymes, optimal activity was observed at pH 7.0 or pH 8.0, and high thermostability was measured up to 55 °C. By contrast, LFFae1432 lost less than 10.0% activity after incubation at 40 °C for 2 h, and pH stability was highest between pH 7.0 and pH 9.0. In addition, LFFae1432 was the most robust esterase, with a higher affinity and hydrolytic activity against synthetic esters. The enzymes released ferulic acids (FAs) from de-starched wheat bran (DSWB), and 60.7% of the total alkali-extractable FAs were released when LFFae1432 was added alone, compared with less than 10% for the other enzymes. The amount of FAs released by FAEs increased when combined with xylanase. These FAEs could serve as promising biocatalysts for biodegradation, and LFFae1432 may hold promise for potential industrial applications. • Thermostable feruloyl esterases (FAEs) from L. fermentum JN248 were screened and identified. • These FAEs presented the higher affinity and hydrolytic activity against synthetic esters. • The yields of ferulic acids by FAEs were increased when combined with xylanase. [ABSTRACT FROM AUTHOR]

Feruloyl esterases (FAEs, E.C. 3.1.1.73) are biotechnologically important enzymes with several applications in ferulic acid production from biomass, but also in synthesis of hydroxycinnamic acid derivatives. The use of such biocatalysts in commercial processes can become feasible by their immobilization, providing the advantages of isolation and recycling. In this work, eight feruloyl esterases, immobilized in cross-linked enzyme aggregates (CLEAs) were tested in regard to their transesterification performance, towards the production of prenyl ferulate (PFA) and arabinose ferulate (AFA). After solvent screening, comparison with the activity of respective soluble enzymes, and operational stability tests, FAE125 was selected as the most promising biocatalyst. A central composite design revealed the optimum conditions for each transesterification product, in terms of water content, time, and substrate ratio for both products, and temperature and enzyme load additionally for prenyl ferulate. The optimum product yields obtained were 83.7% for PFA and 58.1% for AFA. FAE125 CLEAs are stable in the optimum conditions of transesterification reactions, maintaining 70% residual activity after five consecutive reactions. Overall, FAE125 CLEAs seem to be able to perform as a robust biocatalyst, offering satisfactory yields and stability, and thus showing significant potential for industrial applications. [ABSTRACT FROM AUTHOR]

With a production of 39 million metric tons each year, rice is one of the main agricultural products of Vietnam. Thus, rice straw is a significant by-product, whose use in a biorefinery process would contribute to the bio-based transformation of the Vietnamese and South East Asian economy. In order to find novel efficient enzyme mixtures for the hydrolysis of rice straw and other agricultural residues, we took advantage of the rapidly evolving biodiversity of Vietnam and screened 1100 new fungal isolates from soil and decaying plant tissues for their CMCase activity. We selected 36 strains and evaluated them for their cellulases, xylanases, and accessory enzymes activities. Most of these isolates belonged to the genera Aspergillus and Trichoderma . We identified a few promising isolates, such as A. brunneoviolaceus FEC 156, A. niger FEC 130 and FEC 705, and A. tubingensis FEC 98, FEC 110 and FEC 644, whose produced enzyme mixtures released a mass fraction of the sugar content of alkali-treated rice straw higher than 20%, compared to 10% for Trichoderma reesei RUT C-30. We verified that the black Aspergilli are particularly efficient in their saccharification ability. We also identified strains that although they produced low amounts of cellulases and xylanases, their enzyme mixtures had high saccharification efficiencies, indicating the importance of the synergy effect, rather than the amount of enzymes available. Our results highlight the intra-species variation, especially in the Trichoderma genus, regarding the biomass degradation characteristics and the associated range of enzymatic activities. [ABSTRACT FROM AUTHOR]

Feruloyl esterases (Faes) are a subclass of the carboxylic esterases that hydrolyze the ester bonds between ferulic acid and polysaccharides in plant cell walls. Until now, the biochemical characteristics of FAEs from Bacillus spp. have not been reported. In this study, a strain with high activity of FAEs, Bacillus amyloliquefaciens H47 was screened from 122 Bacillus – type strains. Finally, three FAEs (BaFae04, BaFae06, and BaFae09) were identified. Comparing with other bacterial FAEs, these novel FAEs exhibited low sequence identities (less than 30%). The profiles of 52 esterase substrates showed that the three FAEs had a broad substrate spectrum and could effectively hydrolyze several common FAE substrates, such as methyl ferulate, ethyl caffeate, methyl p -coumarate, methyl sinapate, and chlorogenic acid. Furthermore, the three FAEs also can release ferulic acid from destarched wheat bran. They showed maximal activity with an optimal pH of 8.0 at 30 °C, 35 °C, and 40 °C, respectively. BaFae04 showed high stability in the temperature range of 25–60 °C for 1 h and retained 59% of its activity at 60 °C. The present study displays some useful characteristics of FAEs for potential industrial application and contributes to our understanding of FAEs. [ABSTRACT FROM AUTHOR]

Feruloyl esterases (FAEs, E.C. 3.1.1.73) are biotechnologically important enzymes with several applications in ferulic acid production from biomass, but also in synthesis of hydroxycinnamic acid derivatives. The use of such biocatalysts in commercial processes can become feasible by their immobilization, providing the advantages of isolation and recycling. In this work, eight feruloyl esterases, immobilized in cross-linked enzyme aggregates (CLEAs) were tested in regard to their transesterification performance, towards the production of prenyl ferulate (PFA) and arabinose ferulate (AFA). After solvent screening, comparison with the activity of respective soluble enzymes, and operational stability tests, FAE125 was selected as the most promising biocatalyst. A central composite design revealed the optimum conditions for each transesterification product, in terms of water content, time, and substrate ratio for both products, and temperature and enzyme load additionally for prenyl ferulate. The optimum product yields obtained were 83.7% for PFA and 58.1% for AFA. FAE125 CLEAs are stable in the optimum conditions of transesterification reactions, maintaining 70% residual activity after five consecutive reactions. Overall, FAE125 CLEAs seem to be able to perform as a robust biocatalyst, offering satisfactory yields and stability, and thus showing significant potential for industrial applications.

Cross-linked enzyme aggregates (CLEA®) technology is a well-established method in the current literature for the low-cost and effective immobilization of several enzymes. The main advantage of this particular method is the simplicity of the process, since it consists of only two steps. However, CLEA immobilization must be carefully designed for each desired enzyme, since the optimum conditions for enzymes can vary significantly, according to their physicochemical properties. In the present study, an investigation of the optimum CLEA immobilization conditions was carried out for eight feruloyl esterase preparations. Feruloyl esterases are a very important enzyme group in the valorization of lignocellulosic biomass, since they act in a synergistic way with other enzymes for the breakdown of plant biomass. Specifically, we investigated the type and concentration of precipitant and the crosslinker concentration, for retaining optimal activity. FAE68 was found to be the most promising enzyme for CLEA immobilization, since in this case, the maximum retained activity, over 98%, was observed. Subsequently, we examined the operational stability and the stability in organic solvents for the obtained CLEA preparations, as well as their structure. Overall, our results support that the maximum activity retaining and the stability properties of the final CLEAs can vary greatly in different FAE preparations. Nevertheless, some of the examined FAEs show a significant potential for further applications in harsh industrial conditions.

Aims To characterize the hemicellulolytic and cellulolytic enzymes from novel fungi, and evaluate the potential of novel enzyme system in releasing ferulic acid (FA) from biomass resource. Methods and Results A hemicellulolytic and cellulolytic enzyme-producing fungus 4-14 was isolated from soil by Congo red staining method, and identified as Eupenicillium parvum based on the morphologic and molecular phylogenetic analysis. The optimum temperature of fungal growth was 37°C. Hemicellulolytic and cellulolytic enzymes were produced by this fungus in solid-state fermentation ( SSF), and their maximum activities were 554, 385, 218, 2·62 and 5·25 U g−1 for CMCase, xylanase, β-glucosidase, FPase and FAE respectively. These enzymes displayed the best catalytic ability at low pH values ( pH 4·5-5·0). The optimum temperatures were 70°C, 70°C, 75°C and 55°C for CMCase, β-glucosidase, xylanase and FAE respectively. CMCase, xylanase and FAE were stable at different pHs or high temperature (60°C). Enzymatic hydrolysis experiment indicated that the maximum (76·8 ± 4)% of total alkali-extractable FA was released from de-starched wheat bran by the fungal enzyme system. Conclusions High activities of thermotolerant CMCase, β-glucosidase, xylanase and FAE were produced by the newly isolated fungus E. parvum 4-14 in SSF. The fungal enzyme system displayed high efficiency at releasing FA from wheat bran. Significance and Impact of the Study This study provides a new fungal strain for researches of novel hemicellulolytic and cellulolytic enzymes and will improve the bioconversion and utilization of agricultural by-products. [ABSTRACT FROM AUTHOR]

Cosmeceuticals are cosmetic products containing biologically active ingredients purporting to offer a pharmaceutical therapeutic benefit. The active ingredients can be extracted and purified from natural sources (botanicals, herbal extracts, or animals) but can also be obtained biotechnologically by fermentation and cell cultures or by enzymatic synthesis and modification of natural compounds. A cosmeceutical ingredient should possess an attractive property such as anti-oxidant, anti-inflammatory, skin whitening, anti-aging, anti-wrinkling, or photoprotective activity, among others. During the past years, there has been an increased interest on the enzymatic synthesis of bioactive esters and glycosides based on (trans)esterification, (trans)glycosylation, or oxidation reactions. Natural bioactive compounds with exceptional theurapeutic properties and low toxicity may offer a new insight into the design and development of potent and beneficial cosmetics. This review gives an overview of the enzymatic modifications which are performed currently for the synthesis of products with attractive properties for the cosmeceutical industry. [ABSTRACT FROM AUTHOR]

Enzymes' uncharacterised side activities can have significant effects on reaction products and yields. Hence, their identification and characterisation are crucial for the development of successful reaction systems. Here, we report the presence of feruloyl esterase activity in CtXyn5A from Acetivibrio thermocellus, besides its well-known arabinoxylanase activity, for the first time. Activity analysis of enzyme variants mutated in the catalytic nucleophile, Glu279, confirmed removal of all activity for E279A and E279L, and increased esterase activity while removing xylanase activity for E279S, thus allowing the proposal that both reaction types are catalysed in the same active site in two subsequential steps. The ferulic acid substituent is cleaved off first, followed by hydrolysis of the xylan backbone. The esterase activity on complex carbohydrates was found to be higher than that of a designated ferulic acid esterase (E-FAERU). Therefore, we conclude that the enzyme exhibits a dual function rather than an esterase side activity., (© 2022 The Authors. ChemBioChem published by Wiley-VCH GmbH.)

Feruloyl esterase plays an indispensable role in hydrolyzing plant cell walls, and ferulic acid will be released as by-product, which has potential applications in food and medicine industry. This study presents immobilization of partially purified feruloyl esterases from the fermentation liquor of recombinant Pichia on magnetic Fe 3 O 4 nanoparticles. Furthermore, the optimal conditions for the immobilization and some characteristics of immobilized enzyme were studied. The optimal immobilization conditions observed were enzyme 0.2 mg (1 mg/mL, 0.2 mL), magnetic Fe 3 O 4 nanoparticles 4 mg, pH 6.0, immobilization time 3 h. The results showed that the optimal reaction temperature of immobilized enzyme was increased from 45°C to 55°C. Thermal stability of the immobilized enzyme also had an improvement, the residual activity retained 80% and 69% after 120 h at 40°C and 50°C, respectively, while free enzymes only showed 45% and 40% remnant activity at the same condition. The immobilized enzyme also exhibited good operational stability and 52.4% of its initial activity was observed during the fifth cycle. In terms of the thermal and operational stability, the immobilized enzyme could be better used in many more applications than the free enzymes. At last the destarched wheat bran was taken as substrate for immobilized and free feruloyl esterases to produce ferulic acid, the maximum ferulic acid yield was 11.2% and 12.3%, respectively, indicating a great potential in industrial application. [ABSTRACT FROM AUTHOR]

A metagenome expression library was created from Trinervitermes trinervoides termite hindgut symbionts and subsequently screened for feruloyl esterase (FAE) activities, resulting in seven recombinant fosmids conferring feruloyl esterase phenotypes. The amino acid sequence lengths of the seven FAE encoding open reading frames (ORFs) ranged from 260 to 274 aa and encoded polypeptides of between 28.9 and 31.4 kDa. The highest sequence identity scores for the seven ORFs against the GenBank database were between 45 and 59 % to a number of carboxyl ester hydrolyses. The seven FAE primary structures contained sequence motifs that correspond well with a classical pentapeptide (G-x-S-x-G) serine hydrolyse signature motif which harbours the catalytic serine residue in other FAE families. Six of the seven fae genes were successfully expressed heterologously in Escherichia coli, and the purified enzymes exhibited temperature optima range of 40-70 °C and the pH optima of between 6.5 and 8.0. The k/ K ratios for the six characterised FAEs showed the following order of substrate preference: methyl sinapate > methyl ferulate > ethyl ferulate. All six FAEs showed poor conversion rates against methyl p-coumarate and methyl caffeate, both of which lacked the methoxy (O-CH) group substituent on the aromatic ring of the ester substrates, emphasising the requirement for at least one methoxy group on the aromatic ring of the hydroxycinnamic acid ester substrate for optimal FAE activity. [ABSTRACT FROM AUTHOR]

Highlights: [•] Surface charge distribution affects enzyme orientation and substrate accessibility. [•] Immobilization pH alters the specific activity irrespective of reaction pH. [•] Enzyme product selectivity is buffer dependent. [Copyright &y& Elsevier]

Abstract: Feruloyl esterases (FAEs) are a group of important industrial enzymes, which could hydrolyze the ester bonds between hydroxycinnamic acids and arabinose or galactose in hemicellulose present in plant cell walls. To establish a practical high-throughput screening system of feruoyl esterases, two new substrates with modified chromo- or fluoro-phore, 2-chloro-4-nitrophenyl ferulate (CNPF; 1b) and umbelliferyl 5-O-feruolyl-α-L-arabinofuranoside (UFA; 2a), were designed and synthesized. Both substrates provided significantly improved signal-to-noise ratio compared with previously known structural analogues. The chromogenic substrate CNPF could be readily adapted to the high-throughput screening of feruloyl esterase A from Aspergillus niger, one of the most studied FAEs, with the coefficient of variance of as low as 10.3% as determined for a single sequence library. [Copyright &y& Elsevier]

Feruloyl esterases hydrolyze the ester linkages of ferulic and diferulic acids present in plant cell walls. This interesting group of enzymes also has a potentially broad range of applications in the pharmaceutical and agri-food industries. An overview of the current knowledge of fungal feruloyl esterases focusing on the diverse of substrate specificity and potential applications is presented in this review. Furthermore, biological functions of ferulic acid are discussed. [ABSTRACT FROM AUTHOR]

Feruloyl esterase (FAE) and xylanase activities were detected in culture supernatants from Humicola grisea var. thermoidea and Talaromyces stipitatus grown on brewers’ spent grain (BSG) and wheat bran (WB), two agro-industrial by-products. Maximum activities were detected from cultures of H. grisea grown at 150rpm, with 16.9U/ml and 9.1U/ml of xylanase activity on BSG and WB, respectively. Maximum FAE activity was 0.47U/ml and 0.33U/ml on BSG and WB, respectively. Analysis of residual cell wall material after microbial growth shows the preferential solubilisation of arabinoxylan and cellulose, two main polysaccharides present in BSG and WB. The production of low-cost cell-wall-deconstructing enzymes on agro-industrial by-products could lead to the production of low-cost enzymes for use in the valorisation of food processing wastes. [Copyright &y& Elsevier]

Abstract: Commercial multicomponent enzyme preparations, Ultraflo L, Depol 740L and Depol 670L, with feruloyl esterase activity, were tested for the transesterification of methyl ferulate to 1-butyl ferulate in their free and immobilized form using as a reaction system a ternary water–organic mixture consisting of n-hexane, 1-butanol and water. A number of factors affecting enzymes precipitation and cross-linking into cross-linked enzyme aggregates (CLEAs) have been investigated. Consecutive optimization of the precipitant type and cross-linker concentration resulted in CLEAs showing higher operational stability and synthetic activity compared to the free enzymes’ forms. Under certain optimization conditions, conversion yields of 97%, 87% and 5%, were obtained by CLEAs prepared from Ultraflo L, Depol 740L and Depol 670L, respectively. The activities initially present in the three commercial preparations were completely retained after cross-linking resulting in multipurpose biocatalysts which have the potential to carry out different and independent reactions. This work is consistent to the novel CLEA concept called combi-CLEA. [Copyright &y& Elsevier]

Feruloyl esterases are part of the enzymatic spectrum employed by fungi and other microorganisms to degrade plant polysaccharides. They release ferulic acid and other aromatic acids from these polymeric structures and have received an increasing interest in industrial applications such as in the food, pulp and paper and bio-fuel industries. This review provides an overview of the current knowledge on fungal feruloyl esterases focussing in particular on the differences in substrate specificity, regulation of their production, prevalence of these enzymes in fungal genomes and industrial applications. [ABSTRACT FROM AUTHOR]

Abstract: Feruloyl esterases (FAEs) act synergistically with xylanases to hydrolyze ester-linked ferulic (FA) and diferulic (diFA) acid from cell wall material and therefore play a major role in the degradation of plant biomass. The potential applications of these enzymes with reference to agriculture, food and pharmaceutical industries, are discussed in this review. FAE activities produced by different microorganisms are compared for both submerged and solid state fermentations. In addition, their physicochemical properties and molecular biology are presented. [Copyright &y& Elsevier]

Abstract: Fourteen thermophilic and thermotolerant fungal strains isolated from composting soils produced plant cell wall-acting esterases in a medium containing corn cobs and oat spelt xylan. The concentrated and dialyzed protein extracts of these fungi were fractionated using isoelectric-focusing, gels sliced and eluted protein in each slice was assayed for esterase activity against p-nitrophenyl acetate. A total of 84 esterases detected on the basis of pI were found to show distinct preferential substrate specificities towards p-nitrophenyl acetate, p-nitrophenyl ferulate and p-nitrophenyl butyrate, and were putatively classified as acetyl esterases and esterases types I and II. None of the esterases were active against p-nitrophenyl myristate. In addition, these esterases were characterized as acid, neutral or alkaline active. [Copyright &y& Elsevier]

A number of factors affecting production of feruloyl esterase an enzyme that hydrolyse ester linkages of ferulic acid (FA) in plant cell walls, by the thermophylic fungus Sporotrichum thermophile under solid state fermentation (SSF) were investigated. Initial moisture content and type of carbon source were consecutively optimised. SSF in a laboratory horizontal bioreactor using the optimised medium allowed the production of 156 mU g−1 of carbon source, which compared favourably with those reported for the other micro-organisms. Optimal esterase activity was observed at pH 8 and 60 °C. The activity of the esterase was measured on an insoluble feruloylated hemicellulose substrate (de-starched wheat bran (DSWB)). De-esterification of wheat straw yielded loss of feruloyl esterase production even though the supplementation of free FA comparable to the alkali-extractable levels of FA found in wheat straw. Chromogenic (fluorogenic) 4-methylumbelliferyl ferulate was used to characterise the multienzyme component, after separation by isoelectric focusing and native PAGE electrophoresis. The zymograms indicated one major esterase activity exhibiting pI and molecular mass values 5 and 27 kDa, respectively. [Copyright &y& Elsevier]

4-Nitrophenyl 2-O-(E)-feruloyl-α-l-arabinofuranoside 1 and 4-nitrophenyl 5-O-(E)-feruloyl-α-l-arabinofuranoside 2 have been synthesized by two different routes. Monoferuloylation was accomplished by a chemoenzymatic sequence employing a regioselective transesterification catalyzed by lipases. The feruloyl group was introduced to enzymatically prepared 2,3- and 3,5-diacetates of 4-nitrophenyl α-l-arabinofuranoside by reaction with 4-O-acetylferuloyl chloride. Removal of the protecting acetyl groups yielded 1 and 2. An alternative chemical synthesis suitable for preparation of larger quantities of 1 and 2 also is presented. The new substrates represent convenient tools to differentiate feruloyl esterases on the basis of their substrate specificity. [Copyright &y& Elsevier]

Feruloyl esterases (FAEs) have an important role in the enzymatic conversion of lignocellulosic biomass by decoupling plant cell wall polysaccharides and lignin. Moreover, FAEs release anti-oxidative hydroxycinnamic acids (HCAs) from biomass. As a plethora of FAE candidates were found in fungal genomes, FAE classification related to substrate specificity is an indispensability for selection of most suitable candidates. Hence, linking distinct substrate specificities to a FAE classification, such as the recently classified FAE subfamilies (SF), is a promising approach to improve the application of these enzymes for a variety of industrial applications. In total, 14 FAEs that are classified members of SF1, 5, 6, 7, 9, and 13 were tested in this research. All FAEs were investigated for their activity toward a variety of substrates: synthetic model substrates, plant cell wall-derived substrates, including lignin, and natural substrates. Released HCAs were determined using reverse phase-ultra high performance liquid chromatography coupled to UV detection and mass spectrometry. Based on this study, FAEs of SF5 and SF7 showed the highest release of FA, p CA, and diFAs over the range of substrates, while FAEs of SF6 were comparable but less pronounced for diFAs release. These results suggest that SF5 and SF7 FAEs are promising enzymes for biorefinery applications, like the production of biofuels, where a complete degradation of the plant cell wall is desired. In contrast, SF6 FAEs might be of interest for industrial applications that require a high release of only FA and p CA, which are needed as precursors for the production of biochemicals. In contrast, FAEs of SF1, 9 and 13 showed an overall low release of HCAs from plant cell wall-derived and natural substrates. The obtained results substantiate the previous SF classification as a useful tool to predict the substrate specificity of FAEs, which eases the selection of FAE candidates for industrial applications., (Copyright © 2020 Underlin, Frommhagen, Dilokpimol, van Erven, de Vries and Kabel.)

Cross-linked enzyme aggregates (CLEA®) technology is a well-established method in the current literature for the low-cost and effective immobilization of several enzymes. The main advantage of this particular method is the simplicity of the process, since it consists of only two steps. However, CLEA immobilization must be carefully designed for each desired enzyme, since the optimum conditions for enzymes can vary significantly, according to their physicochemical properties. In the present study, an investigation of the optimum CLEA immobilization conditions was carried out for eight feruloyl esterase preparations. Feruloyl esterases are a very important enzyme group in the valorization of lignocellulosic biomass, since they act in a synergistic way with other enzymes for the breakdown of plant biomass. Specifically, we investigated the type and concentration of precipitant and the crosslinker concentration, for retaining optimal activity. FAE68 was found to be the most promising enzyme for CLEA immobilization, since in this case, the maximum retained activity, over 98%, was observed. Subsequently, we examined the operational stability and the stability in organic solvents for the obtained CLEA preparations, as well as their structure. Overall, our results support that the maximum activity retaining and the stability properties of the final CLEAs can vary greatly in different FAE preparations. Nevertheless, some of the examined FAEs show a significant potential for further applications in harsh industrial conditions. [ABSTRACT FROM AUTHOR]

Carbohydrate esterases are a group of enzymes which release acyl or alkyl groups attached by ester linkage to carbohydrates. The CAZy database, which classifies enzymes that assemble, modify, and break down carbohydrates and glycoconjugates, classifies all carbohydrate esterases into 16 families. This chapter is an overview of the research for nearly 50 years around the main groups of carbohydrate esterases dealing with the degradation of polysaccharides, their main biochemical and molecular traits, as well as its application for the synthesis of high added value esters.