Enzymatic Hydrolysis Performance Research Articles

Structural change in wood by brown rot fungi and effect on enzymatic hydrolysis

The effects of biological pretreatment on Pinus radiata and Eucalyptus globulus, were evaluated after exposure to two brown rot fungi Gloephylum trabeum and Laetoporeus sulphureus. Changes in chemical composition, structural modification, and susceptibility to enzymatic hydrolysis in the degraded wood were analyzed. After eight weeks of biodegradation, the greatest loss of weight and hemicellulose were 13% and 31%, respectively, for P. radiata with G. trabeum. The content of glucan decreased slightly, being the highest loss of 20% for E. globulus with G. trabeum. Consistent with degradation mechanism of these fungi, lignin was essentially undegraded by both brown rot fungi. Both brown rot fungi cause a sharp reduction in the cellulose degree of polymerization (DP) in the range between 58% and 79%. G. trabeum depolymerized cellulose in both wood faster than L. sulphureus. Also, structural characteristic of crystalline cellulose were measured by using two different techniques – X-ray diffraction (XRD) and infrared spectroscopy (FT-IR). The biological pretreatments showed an effect on cellulose crystallinity structure, a decrease between 6% and 21% was obtained in the crystallinity index (CrI) calculated by IR, no changes were observed in the XRD. Material digestibility was evaluated by enzymatic hydrolysis, the conversion of cellulose to glucose increased with the biotreatment time. The highest enzymatic hydrolysis yields were obtained when saccharification was performed on wood biopretreated with G. trabeum (14% P. radiata and 13% E. globulus). Decreasing in DP and CrI, and hemicellulose removal result in an increase of enzymatic hydrolysis performance. Digestibility was better related to DP than with other properties. G. trabeum can be considered as a potential fungus for biological pretreatment, since it provides an effective process in breaking the wood structure, making it potentially useful in the development of combined pretreatments (biological–chemical). A viable alternative to pretreatment process that can be used is a bio-mimetic system, similar to low-molecular complexes generated by fungi such as G. trabeum combined pretreatments (biological–chemical).

Particle morphology characterization and manipulation in biomass slurries and the effect on rheological properties and enzymatic conversion

An improved understanding of how particle size distribution relates to enzymatic hydrolysis performance and rheological properties could enable enhanced biochemical conversion of lignocellulosic feedstocks. Particle size distribution can change as a result of either physical or chemical manipulation of a biomass sample. In this study, we employed image processing techniques to measure slurry particle size distribution and validated the results by showing that they are comparable to those from laser diffraction and sieving. Particle size and chemical changes of biomass slurries were manipulated independently and the resulting yield stress and enzymatic digestibility of slurries with different size distributions were measured. Interestingly, reducing particle size by mechanical means from about 1 mm to 100 μm did not reduce the yield stress of the slurries over a broad range of concentrations or increase the digestibility of the biomass over the range of size reduction studied here. This is in stark contrast to the increase in digestibility and decrease in yield stress when particle size is reduced by dilute-acid pretreatment over similar size ranges.

PENINGKATAN UNJUK KERJA HIDROLISIS ENZIMATIK JERAMI PADI MENGGUNAKAN CAMPURAN SELULASE KASAR DARI Trichoderma reesei DAN Aspergillus niger

Increasing the Performance of Enzymatic Hydrolysis of Rice Straw Using Mixed Crude Cellulases from Trichoderma reesei and Aspergillus niger. The objective of this work is to compare the effectiveness of mixed crude enzyme cellulase from T. reesei and A. niger with commercial enzyme from A. niger, and to investigate effect of enzyme to substrate ratio to performance of enzymatic hydrolysis of rice straw. The commercial enzyme from Fluka Biochemica was used, and crude enzyme were prepared by solid fermentation with simple media. Before hydrolized, the rice straw was grinded and sieved and then heated at 85 oC with 2% sodium hydroxide for six hours. Hydrolysis was conducted in 300 mL beaker flask equipped with mechanical stirrer. Samples were analyzed by dinitrosalicylic acid method and measured by spectrophotometer. Both of commercial and mixed crude enzyme show that, the higher enzyme to substrate ratio was higher the glucose concentration obtained. However, ratio of glucose obtained to enzyme used become smaller. The mixture of crude enzyme from T. reesesi dan A. niger that produced in this work was two fold more effective to hydrolyze rice straw than using cellulase enzyme of A. niger from Fluka Biochemika. Keywords: rice straw, cellulase, hydrolysis, A. niger, T. reesei

Enzymatic hydrolysis of sugarcane bagasse pretreated with acid or alkali

The aim of this study was to evaluate the performance of enzymatic hydrolysis of acid or alkali pretreated sugarcane bagasse for the production of fermentable sugars. The first step consisted of selection of commercial enzymes presenting the highest cellulolytic activities. After selection of four enzymes: HPL, CL, P1 and P4, their performances were tested in the bagasse pretreated with acid and alkali. The sugar content of the hydrolysates was analyzed by anion exchange liquid chromatography. Data showed that the joint action of 0.5% acid pretreatment, 121ºC, 30 minutes and enzyme CL provides the best results, 67.25 g of hexose and 148.13g of pentose per kg of dry bagasse.

Comparing biological and thermochemical processing of sugarcane bagasse: An energy balance perspective

The technical performance of lignocellulosic enzymatic hydrolysis and fermentation versus pyrolysis processes for sugarcane bagasse was evaluated, based on currently available technology. Process models were developed for bioethanol production from sugarcane bagasse using three different pretreatment methods, i.e. dilute acid, liquid hot water and steam explosion, at various solid concentrations. Two pyrolysis processes, namely fast pyrolysis and vacuum pyrolysis, were considered as alternatives to biological processing for the production of biofuels from sugarcane bagasse. For bioethanol production, a minimum of 30% solids in the pretreatment reactor was required to render the process energy self-sufficient, which led to a total process energy demand equivalent to roughly 40% of the feedstock higher heating value. Both vacuum pyrolysis and fast pyrolysis could be operated as energy self-sufficient if 45% of the produced char from fast pyrolysis is used to fuel the process. No char energy is required to fuel the vacuum pyrolysis process due to lower process energy demands (17% compared to 28% of the feedstock higher heating value). The process models indicated that effective process heat integration can result in a 10–15% increase in all process energy efficiencies. Process thermal efficiencies between 52 and 56% were obtained for bioethanol production at pretreatment solids at 30% and 50%, respectively, while the efficiencies were 70% for both pyrolysis processes. The liquid fuel energy efficiency of the best bioethanol process is 41%, while that of crude bio-oil production before upgrading is 67% and 56% via fast and vacuum pyrolysis, respectively. Efficiencies for pyrolysis processes are expected to decrease by up to 15% should upgrade to a transportation fuel of equivalent quality to bioethanol be taken into consideration.

Enhanced enzymatic hydrolysis of rapeseed straw by popping pretreatment for bioethanol production

The objective of this study was to find a pretreatment process that enhances enzymatic conversion of biomass to sugars. Rapeseed straw was pretreated by two processes: a wet process involving wet milling plus a popping treatment, and a dry process involving popping plus dry milling. The effects of the pretreatments were studied both in terms of structural and compositional changes and change in susceptibility to enzymatic hydrolysis. After application of the wet and dry processes, the amounts of cellulose and xylose in the straw were 37–38% and 14–15%, respectively, compared to 31% and 12% in untreated counterparts. In enzymatic hydrolysis performance, the wet process presented the best glucose yield, with a 93.1% conversion, while the dry process yielded 69.6%, and the un-pretreated process yielded <20%. Electron microscopic studies of the straw also showed a relative increase in susceptibility to enzymatic hydrolysis with pretreatment.

Engineering of a high‐throughput screening system to identify cellulosic biomass, pretreatments, and enzyme formulations that enhance sugar release

The recalcitrance of cellulosic biomass, the only abundant, sustainable feedstock for making liquid fuels, is a primary obstacle to low cost biological processing, and development of more easily converted plants and more effective enzymes would be of great benefit. Because no single parameter describes recalcitrance, superior variants can only be identified by measuring sugar release from plants subjected to pretreatment and enzymatic hydrolysis. However, genetic modifications of plants coupled with molecular engineering of deconstruction proteins and definition of pretreatment conditions create a very large sample set, and previous methods for biomass pretreatment at elevated temperatures and pressures prevented use of a fully integrated high-throughput (HTP) screening pipeline. Herein, we report on the engineering of a novel HTP pretreatment system employing a 96 well-plate format that withstands extreme pretreatment conditions for rapid screening of biomass-enzyme-pretreatment combinations. This includes the development of new approaches to steam heating and water quenching the system that result in much faster heat up and cool down than previously possible and show consistent temperature histories across the multiwell plate. Coupled pretreatment and enzymatic hydrolysis performance of the well plate pretreatment system is shown to be consistent among the many wells in the device and also with performance of conventional tubular reactors.

난황 단백질의 효소 가수분해에 의한 sialic acid의 회수 공정 개발

난황 단백질의 효소에 의한 가수분해에서 에탄올의 농도가 증가할수록 고형물과 단백질의 회수가 증가함을 알 수 있었다. 이것은 유화층을 에탄올의 증가가 감소시켰기 때문이다. 그러나 가수분해물에서 단백질 함량이나 sialic acid의 함량은 에탄을 농도와는 관계없이 일정하였다. 한외여과 후에 retentate에 대한 에탄올의 영향을 조사하였다. 마찬가지로 고형물의 회수는 에탄을 농도의 증가와 함께 증가하였다. 그리고 retentate에서 sialic acid의 함량은 대략 <TEX>$2.5\%$</TEX>정도로 일정하고 에탄올의 농도에 영향을 받지 않았다 이상의 결과로부터 난황 단백질의 효소 가수분해를 통해 sialic acid가 함유된 제품을 얻고자 할 때는 원료 난황단백질에 포함된 에탄올의 함량을 증가시킬수록 높은 수율의 제품을 얻을 수 있다. 본 실험에서는 원료 난황 단백질 중에서 <TEX>$40\%$</TEX>의 에탄을 함량까지는 제품 수율이 계속 증가하는 경향을 보였다. 난황 단백질 가수분해물의 한외여과에서 농축단계에서는 막 모듈의 MWCO의 차이에도 불구하고 retentate에서 총 고형물의 회수율은 비슷함을 나타내었으며, 투석에서 회수율은 MWCO가 작을수록 높아지지는 않았다. 제품에서의 sialic acid의 함량은 사용한 모듈에서 약 <TEX>$2.0\%$</TEX>를 나타내었다. 이것은 단백질 가수분해물에 비해 5배정도 상승한 결과이다. 본 연구에서 사용된 막 모듈 가운데서 Amicon 모듈이 제품의 회수율과 함량면에서 가장 우수한 특성을 보였다. 결론적으로 난황 단백질의 protease에 의한 가수분해에서 한외여과에 의해 순도를 높일 때 MWCO, 막 모듈의 type 그리고 운전조건 등을 잘 고려해 줄 때 최적의 조업조건을 얻을 수 있었다. Batch enzymatic hydrolysis of egg yolk protein by protease was carried out at laboratory scale coupled to an ultrafiltration module. Effect of ethanol concentrations on the performance of enzymatic hydrolysis was studied to determine the optimum condition of recovery of hydrolysate. The enzymatic hydrolysis was conducted stepwise with following conditions, <TEX>$50^{\circ}C$</TEX>, pH 10.0 and pH 6.5. Ethanol concentration was changed from 10 to <TEX>$40\%$</TEX> (w/w). As ethanol concentration was increased, the recovery yield of total solid and protein in enzymatic hydrolysate was also increased. The content of sialic acid and protein in hydrolysate was independent of ethanol concentration. We also investigated the effect of ethanol concentration on the performance of ultrafiltration. As the concentration of ethanol in yolk protein was increased, the recovery yield of product was increased. Ultra­filtration of egg yolk protein hydrolysate was conducted to increase the content of sialic acid. Four ultrafiltation modules were used in this study, and we evaluated the performance of the UF modules. When Amicon module was used, the recovery percentage of total solid in retentate was <TEX>$6.0\%$</TEX>, which is the highest among the modules used. In spite of the difference in the recovery yield of total solid, the purity of sialic acid in retentate was about <TEX>$2.0\%$</TEX>, which was 5 times higher than that in feed. It was concluded that the recovery yield and the purity of sialic acid did not correlate with the types of modules and the size of MWCO.

Large-scale enzymatic hydrolysis of agricultural lignocellulosic biomass. Part 1: Pretreatment procedures

A process for conversion of lignocellulosic biomass into acetone-butanol (ABE) involving steam explosion pretreatment of the raw material, cellulase production, enzymatic hydrolysis and acetone-butanol fermentation of hydrolysates has been developed. Procedures and scale-up of the steam-explosion pretreatment of corncobs are described. The influence of experimental parameters, time, temperature and addition of acids, on both the performance of enzymatic hydrolysis and the fermentability into ABE of the hydrolysates obtained were first studied in a batch pilot reactor. The same criteria of performance were applied in large scale development work which was completed using a continuous industrial plant (Stake II machine with a capacity of 2–4 t per h) located in the biomass conversion facilities at Soustons, France. Under optimal conditions, which were strongly dependent on the technology used, nearly quantitative hydrolysis yields could be obtained with moderate amounts of appropriate cellulase preparations.

Determination of PCMX Conjugated Metabolites in Blood Plasma and Urine Using Enzymatic Hydrolysis and High Performance Liquid Chromatography

Abstract An enzymatic-high performance liquid chromatography method with electrochemical detection was developed for the quantitation of p-chloro-m-xylenol conjugated metabolites in blood plasma and urine. The method is based on enzymatic hydrolysis of PCMX conjugates and extraction with benzene. The HPLC system consisted of a C18 reverse-phase column and a 65% methanol:35% ammonium carbonate 0.05% solution as a mobile phase. An electrochemical detector/glassy carbon electrode set at a potential of +0.9V versus Ag/AgCl/3M NaCl was used to monitor the drug. The recovery of PCMX after enzymatic hydrolysis from blood plasma samples and urine samples is approximately 98%.