Optimum Conditions For Enzymatic Hydrolysis Research Articles

Optimizing the Conditions of Pretreatment and Enzymatic Hydrolysis of Sugarcane Bagasse for Bioethanol Production.

The agricultural waste sugarcane bagasse (SCB) is a kind of plentiful biomass resource. In this study, different pretreatment methods (NaOH, H2SO4, and sodium percarbonate/glycerol) were utilized and compared. Among the three pretreatment methods, NaOH pretreatment was the most optimal method. Response surface methodology (RSM) was utilized to optimize NaOH pretreatment conditions. After optimization by RSM, the solid yield and lignin removal were 54.60 and 82.30% under the treatment of 1% NaOH, a time of 60 min, and a solid-to-liquid ratio of 1:15, respectively. Then, the enzymolysis conditions of cellulase for NaOH-treated SCB were optimized by RSM. Under the optimal enzymatic hydrolysis conditions (an enzyme dose of 18 FPU/g, a time of 64 h, and a solid-to-liquid ratio of 1:30), the actual yield of reducing sugar in the enzyme-treated hydrolysate was 443.52 mg/g SCB with a cellulose conversion rate of 85.33%. A bacterium, namely, Bacillus sp. EtOH, which produced ethanol and Baijiu aroma substances, was isolated from the high-temperature Daqu of Danquan Baijiu in our previous study. At last, when the strain EtOH was cultured for 36 h in a fermentation medium (reducing sugar from cellulase-treated SCB hydrolysate, yeast extract, and peptone), ethanol concentration reached 2.769 g/L (0.353%, v/v). The sugar-to-ethanol and SCB-to-ethanol yields were 13.85 and 11.81% in this study, respectively. In brief, after NaOH pretreatment, 1 g of original SCB produced 0.5460 g of NaOH-treated SCB. Then, after the enzymatic hydrolysis, reducing sugar yield (443.52 mg/g SCB) was obtained. Our study provided a suitable method for bioethanol production from SCB, which achieved efficient resource utilization of agricultural waste SCB.

Pretreatment and enzymatic hydrolysis optimization of lignocellulosic biomass for ethanol, xylitol, and phenylacetylcarbinol co-production using Candida magnoliae.

Cellulosic bioethanol production generally has a higher operating cost due to relatively expensive pretreatment strategies and low efficiency of enzymatic hydrolysis. The production of other high-value chemicals such as xylitol and phenylacetylcarbinol (PAC) is, thus, necessary to offset the cost and promote economic viability. The optimal conditions of diluted sulfuric acid pretreatment under boiling water at 95°C and subsequent enzymatic hydrolysis steps for sugarcane bagasse (SCB), rice straw (RS), and corn cob (CC) were optimized using the response surface methodology via a central composite design to simplify the process on the large-scale production. The optimal pretreatment conditions (diluted sulfuric acid concentration (% w/v), treatment time (min)) for SCB (3.36, 113), RS (3.77, 109), and CC (3.89, 112) and the optimal enzymatic hydrolysis conditions (pretreated solid concentration (% w/v), hydrolysis time (h)) for SCB (12.1, 93), RS (10.9, 61), and CC (12.0, 90) were achieved. CC xylose-rich and CC glucose-rich hydrolysates obtained from the respective optimal condition of pretreatment and enzymatic hydrolysis steps were used for xylitol and ethanol production. The statistically significant highest (p ≤ 0.05) xylitol and ethanol yields were 65% ± 1% and 86% ± 2% using Candida magnoliae TISTR 5664. C. magnoliae could statistically significantly degrade (p ≤ 0.05) the inhibitors previously formed during the pretreatment step, including up to 97% w/w hydroxymethylfurfural, 76% w/w furfural, and completely degraded acetic acid during the xylitol production. This study was the first report using the mixed whole cells harvested from xylitol and ethanol production as a biocatalyst in PAC biotransformation under a two-phase emulsion system (vegetable oil/1M phosphate (Pi) buffer). PAC concentration could be improved by 2-fold compared to a single-phase emulsion system using only 1M Pi buffer.

Optimization of Enzymatic Hydrolysis of Mackerel (Rastrelliger sp.) Muscle Protein Hydrolysate Using Response Surface Methodology

Mackerel (Rastrelliger sp.) is a widely distributed epipelagic species in South East Asia. Mackerel has a high amount nutrient such as protein (20.83 %) and fat (1.03 %). The high amount of protein and low amount of fat will allow it to be used as a material to produce a good protein hydrolysate. The aim of this study is to determine the optimal enzymatic hydrolysis conditions (time, temperature, and pH) using Response Surface Methodology (RSM). Mackerel Protein Hydrolysate (MPH) was prepared using commercial Flavourzyme. Optimization of MPH was performed by employing Box Behnken Design method of RSM. SN-TCA method was used to calculate the degree of hydrolysis (DH) which is the key parameter in hydrolysis reaction. Optimum hydrolysis conditions were obtained at pH 7, temperature 55°C and 60 min of process. Under these conditions the DH obtained was 17.7293 % with 4% enzyme to substrate ratio. The suggested model for the hydrolysis process is quadratic with the desirability factor of 1. The MPH was further assessed for its amino acid composition using High Performance Liquid Chromatography (HPLC). The hydrolysis process increases the amino acid amounts namely L-Glutamic Acid (19.77%), L-Valin (14.20%), L-Aspartic Acid (11.42%), Glycine (11.04%), L-Alanin (14.20%), L-Prolin (16.80%), and L-Histidin (27.06%). The study suggested that mackerel muscle can be considered to be utilized as fish protein hydrolysis materials.

Investigation into Optimal Conditions for Enzymatic Hydrolysis of Cassava Starch to Glucose by Amylase from Rice

The challenge of finding locally available materials in abundance to meet up with the increase in demand for glucose syrup necessitated this study. Enzymatic hydrolysis of cassava starch to glucose using glucose amylase sourced from rice was conducted. A-3 factor with 6 levels factorial viz. substrate concentration (0.5, 1.0, 1.5, 2.0, 2.5 and 3.0% w/v), pH (4, 5, 6, 7, 8 and 9) and temperature (30, 40, 50, 60, 70 and 80 °C) experiment were employed. Rice malt was prepared and enzyme activated. Starch (substrate), buffer solutions, standard glucose solution and its calibration curve were also prepared. Starch was hydrolyzed by α-amylase and tested for presence of reducing sugar using Benedict solution. Time course of the reaction was studied and enzyme activity determined. It was observed that as reaction time increased (t), amount of glucose produced [P] initially increased but soon recorded infinitesimal increase and later assumed constant. The effects of substrate concentration, pH and temperature were found to be essential on glucose production. Statistical analysis on the effect of substrate concentration [S], reaction time and their interactions showed significant impact at probability level of (p) = 0.05.

Antioxidant activity of bioactive peptides from peanuts protein hydrolysate

Peanut protein hydrolysate was produced from peanut protein isolate hydrolyzed using Alcalase® 2.4 L. Hydrolysis conditions were optimized using a response surface methodology. The two independent variables were the substrate/enzyme ratio and the hydrolysis time with lower and upper limits of (20:1-50:1 v/v) and (1-3 hours), respectively. Both were designed at three levels of central composite design from the response surface methodology and the degree of hydrolysis as the dependent variable. This study aimed to obtain the optimum conditions for enzymatic hydrolysis of peanut protein isolate using Alcalase to produce protein hydrolysate with the maximum degree of hydrolysis. The optimum hydrolysis conditions obtained were the substrate/enzyme ratio of 20:1 (v/v) and the hydrolysis time of 3 hours with a predicted degree of hydrolysis of 40.54%. The actual experimental results obtained a degree of hydrolysis of 40.35% (p>0.05), which showed no difference between the predicted and experimental results. The highest antioxidant activity was obtained from the combination of these treatments, indicated by IC50 of 5.61 ppm.

Enzymatic hydrolysis and detection of 3‐monochloropropane‐1,2‐diol esters

AbstractThere are more and more studies on the detection method of 3‐chloro‐1,2‐propanediol fatty acid esters (3‐MCPD esters) at present, by comparing these methods for the determination of 3‐MCPD esters. Indirect methods, which determine total amount of 3‐MCPD after hydrolysis of the esters, have an advantage over direct methods. The existing indirect methods, however, may yield unreliable results or require long hours of alkaline methanolysis. In contrast, the indirect enzymatic hydrolysis method has mild conditions and more accurate results. In this study, we developed a reliable and rapid indirect method for determination of 3‐MCPD esters. 3‐MCPD esters were enzymolysis to 3‐MCPD by indirect enzymatic hydrolysis method, and the conditions of enzymatic hydrolysis were optimized, the content of 3‐MCPD after enzymatic hydrolysis was detected by gas chromatography–mass spectrometry (GC–MS) and the yield was calculated. Finally, the optimum conditions for enzymatic hydrolysis of 3‐MCPD esters were determined. According to the optimal enzymatic hydrolysis condition, the contents of 3‐MCPD esters in four food oils were determined. The method is simple and sensitive and can meet the requirement of 3‐MCPD esters detection in general oils.

Silver Carp (Hypophthalmichthys molitrix) Scales Collagen Peptides (SCPs): Preparation, Whitening Activity Screening and Characterization

This study involves the preparation of scale collagen peptides (SCPs) with whitening activity from silver carp (Hypophthalmichthys molitrix) and their characterization and peptide sequence identification. In this article, scanning electron microscopy (SEM) was used to observe structure changes of sliver carp scales; enzymatic hydrolysis was optimized through protease screening and response surface optimization. The ultrafiltration was used to separate SCPs and the whitening activity was comprehensively evaluated using radical scavenging rate and tyrosinase-inhibiting activity, among others. An optimal component was characterized and identified using various modern spectral analysis techniques. The results showed that the surface of silver carp scales after decalcification was smooth and clear. The pepsin had the highest peptide yield and tyrosinase-inhibiting activity (90.01% and 82.25%, respectively). The optimal enzymatic hydrolysis conditions were an enzyme dosage of 16.1%, a solid-liquid ratio of 1:15.6 and a time of 4.9 h. The proportions of hydrophobic and basic amino acids in the peptide composition were 32.15% and 13.12%, respectively. Compared with SCPs2, SCPs1 (6096.68-9513.70 Da) showed better ·OH scavenging ability, tyrosinase-inhibiting activity and moisture absorption. SCPs1 was a macromolecular fragment of type I collagen with a triple helix structure, containing three peptide sequences with the potential for tyrosinase activity inhibition (AGPPGADGQTGQRGE, SGPAGIAGPAGPRGPAGPNGPPGKD and KRGSTGEQGSTGPLGMRGPRGAA). These results show that SCPs1 is a collagen peptide product with whitening potential.

The Preparation and Physiochemical Characterization of Tenebrio molitor Chitin Using Alcalase.

Chitin is mostly produced from crustaceans, but it is difficult to supply raw materials due to marine pollution, and the commonly used chemical chitin extraction method is not environmentally friendly. Therefore, this study aims to establish a chitin extraction process using enzymes and to develop edible insect-derived chitin as an eco-friendly new material. The response surface methodology (RSM) was used to determine the optimal conditions for enzymatic hydrolysis. The optimal conditions for enzymatic hydrolysis by RSM were determined to be the substrate concentration (7.5%), enzyme concentration (80 μL/g), and reaction time (24 h). The solubility and DDA of the mealworm chitosan were 45% and 37%, respectively, and those of the commercial chitosan were 61% and 57%, respectively. In regard to the thermodynamic properties, the exothermic peak of mealworm chitin was similar to that of commercial chitin. In the FT-IR spectrum, a band was observed in mealworm chitin corresponding to the C=O of the NHCOCH3 group at 1645 cm-1, but this band showed low-intensity C=O in the mealworm chitosan due to deacetylation. Collectively, mealworm chitosan shows almost similar physical and chemical properties to commercial chitosan. Therefore, it is shown that an eco-friendly process can be introduced into chitosan production by using enzyme-extracted mealworms for chitin/chitosan production.

Effective fractionation of lignocellulose components and lignin valorization by combination of deep eutectic solvent with ethanol.

Introduction: A combination of deep eutectic solvent with ethanol was developed for pretreatment of Broussonetia papyrifera to effectively extract lignin and promote the subsequent enzymatic hydrolysis. Methods: In order to further explore the optimal conditions for enzymatic hydrolysis, a central composite design method was applied. Results and Discussion: The correlation between each factor and glucose yield was obtained, and the optimal conditions was 160°C, 60min, the ratio of DES to E was 1/1 (mol/mol). The results showed that compared with control, the glucose yield increased by 130.67% under the optimal pretreatment conditions. Furthermore, the specific surface area of biomass was increased by 66.95%, and the content of xylan and lignin was decreased by 86.71% and 85.83%. The correlation between xylan/lignin removal and enzymatic hydrolysis showed that the removal of lignin facilitated the glucose yield more significantly than that of xylan. To further explore the lignin valorization, the structural and antioxidant analysis of recovered lignin revealed that high temperature was favorable for lignin with good antioxidant performance. This pretreatment is a promising method for separating lignin with high antioxidant activity and improving cellulose digestibility.

Production of Xylooligosaccharides from Jiuzao by Autohydrolysis Coupled with Enzymatic Hydrolysis Using a Thermostable Xylanase.

The production of xylooligosaccharides (XOS) from Jiuzao was studied using a two-stage process based on autohydrolysis pretreatment followed by enzymatic hydrolysis. Jiuzao was autohydrolyzed under conditions where temperature, time, particle size, and solid-liquid ratio were varied experimentally. Optimal XOS production was obtained from Jiuzao with a >20 mesh particle size treated at 181.5 °C for 20 min with a 1:13.6 solid-liquid ratio. Subsequently, optimal enzymatic hydrolysis conditions for xylanase XynAR were identified as 60 °C, pH 5, and xylanase XynAR loading of 15 U/mL. Using these conditions, a yield of 34.2% XOS was obtained from Jiuzao within 2 h. The process developed in the present study could enable effective and ecofriendly industrial production of XOS from Jiuzao.

Ultrasonic‐assisted extraction and purification of xylo‐oligosaccharides from wheat bran

AbstractThis study was to produce functional xylo‐oligosaccharides (XOS) from wheat bran via ultrasonic‐assisted extraction, enzymatic hydrolysis, and purification. Optimal extraction conditions were ultrasonic frequency of 72 kHz, sonication time of 20 min, solid/liquid ratio of 1:21(w/v), 100°C, and extraction time of 2.25 h. Under these conditions, XOS extraction rate of 60.27% was obtained. Optimal enzymatic hydrolysis conditions were 10% for substrate concentration, 800 U/g for enzyme concentration, 55°C, and 4 h for hydrolysis time. Under these conditions, a hydrolysate with 25.52% of XOS content, DP = 2.68. In addition, the hydrolysate solution was decolorized using anion‐exchange resin D392 and the highest decolorization rate (89.26%) was obtained at elution volume of 1 ml/min and 50°C. Decolorized XOS hydrolysate was purified using cation‐exchange resin under optimized conditions of 1.5 ml/min elution volume, 11 ml injection volume, and 35% concentration at 75°C. The concentration of XOS increased from 64.41% to 75.63% after purification.

Expeller-Pressed Pomegranate Seed (Punica granatum L.) as a Protein Source for the Production of Antioxidant Peptides

Because of health concerns regarding synthetic antioxidants, natural antioxidant compounds are being considered by scientists. Bioactive peptides have been shown to have various physiological functions, such as antioxidative activity. Pomegranate seed protein, the by-product of the pomegranate seed oil industry, can be a good source of bioactive peptides. The optimum conditions for enzymatic hydrolysis of pomegranate seed protein with alcalase were determined using a response surface methodology. The influence of different temperatures (45–55 °C), times (30–180 min), and enzyme to substrate (E/S) ratio (1–3% w/w) on DPPH scavenging power and ferric reducing activity as the responses, were studied. Also, the degree of hydrolysis and the surface hydrophobicity of samples were determined. Moreover, using scanning electron microscopy and electrophoresis technique, microscopic structure and molecular weight of hydrolysate, were studied respectively. Alcalase-derived hydrolysates showed a DPPH scavenging activity (88 ± 0.97%) and ferric reducing power (0.5 ± 0.83) at optimum conditions of hydrolysis (48.8 °C, 97.5 min, E/S ratio 1.3%w/w). The degree of hydrolysis coincided with 36 ± 1.2%. An increase in the surface hydrophobicity of the protein during hydrolysis confirmed the unfolding of the pomegranate seed protein structure. The presence of low-molecular-weight peptides was evidenced by the electrophoresis technique. As well as the SEM showed that protein fragments had been reduced to small sizes following enzymatic treatments. According to the results of this study, pomegranate seed protein hydrolysate can be considered a suitable source of antioxidants with an aggregate market value in food formulations.

Investigation of lactic acid production by pressurized liquid hot water from cultivated Miscanthus × giganteus

The production of lactic acid, a polylactic acid monomer from energy crop Miscanthus × giganteus lignocellulosic biomass cultivated in Izmir was investigated. Liquid hot water (LHW) pretreatment was carried out at a temperature range of 140–200 °C, pressure 100 to 200 bar and reaction time of 15–45 min at a fixed flow rate of 2 mL/min using D-optimal experimental plan. The optimum conditions were elicited as 140 °C, 100 bar and 45 minutes, yielding the highest reducing sugar content of 77.32 mg/g, whereas 1.25 mg/mL arabinose and 1.35 mg/mL xylose as monomeric sugars. Subsequently, the enzymatic hydrolysis was applied to the solid fraction. The optimum conditions for enzymatic hydrolysis were determined as 5% (w/v) solid/liquid ratio, 20 FPU/mL enzyme loading and 72 hours, revealing the highest amount of reducing sugar as 200 mg/mL. LHW hydrolysate was used as a production medium for lactic acid manufacturing in submerged fermentation by Rhizopus oryzae. The maximum lactic acid content was found to be 6.8 g/L at 24 hours, whereas the lactic acid yield was 0.28 g/L.h. The sequential design of LHW, followed by enzymatic hydrolysis and submerged lactic acid fermentation can be utilized in industry, contributing to the bioeconomy.

Preparation of angiotensin I-converting enzyme (ACE) inhibitory peptides from Tie Guanyin tea residue protein using two-step enzymatic hydrolysis

In this study, we attempted to prepare novel angiotensin I-converting enzyme inhibitory (ACEI) peptides from Tie Guanyin tea residue protein (TTRP). Different proteases were used to hydrolyze TTRP to prepare ACEI peptides, neutral protease and alkaline protease hydrolysates exhibited higher ACEI activity and was chosen in further hydrolysis treatment. Response surface methodology (RSM) was used to optimize the enzymatic condition, and the results indicated that the optimal conditions of enzymatic hydrolysis were: 4% (w/v) of TTRP in water was hydrolyzed with 3500 U/g neutral protease at 50 °C and pH 6.5 for 2.5 h in the first step and then with 5000 U/g alkaline protease at 50 °C and pH 10.0 for 3 h in the second step. Under the optimum condition, the ACE inhibitory rate of the final products was 88.45% ± 0.45% at the concentration of 1.0 mg/mL. Moreover, ACEI peptides derived from TTRP showed good stability under different temperatures, pHs, metal ions and gastrointestinal digestion. These results indicated that the TTRP has potential to be used for preparing ACEI peptides.

Optimization of Enzymatic Preparation Process of Hypoglycemic Peptide from Moringa oleifera Seeds by Response Surface Methodology and Its Hypoglycemic Activities Evaluation in Vitro

In order to improve the resource utilization of Moringa oleifera seeds powder, enzymatic method was used to prepare hypoglycemic peptide from Moringa oleifera seeds. Taking the the degree of proteolysis and α-glucosidase inhibition rate as the response value, single-factor experimental screening and response surface method was used to optimize the optimum enzymatic hydrolysis time, liquid-to-material ratio, hydrolysis pH, enzyme addition, and enzymatic hydrolysis temperature. The hypoglycemic peptide with a molecular weight of <3 kDa were prepared by coarse separation by ultrafiltration, and their inhibitory effects on human liver cancer cells (HepG2 cells) were analyzed by MTT method. The results showed that the optimal conditions of enzymatic hydrolysis were as follows: Time 4.6 h, ratio of liquid to material 40.5:1, pH8.3, enzyme dosage 5.5%, temperature 55 ℃, and the inhibition rate of α-glucosidase was 23.62%±0.14%. The α-glucosidase inhibition rate of the ultrafiltration fraction with a molecular weight of less than 3 kDa had an IC50 value of 5.56 mg/mL. When the concentration of ultrafiltration fraction was 300 μg/mL, the proliferation of HepG2 cells was significantly inhibited after 48 h(P<0.05). The research can lay the foundation for the further separation and purification of hypoglycemic peptide from Moringa oleifera seeds.

Optimisation of enzymatic saccharification of wheat straw pre-treated with sodium hydroxide

To enhance the reducing sugar yield in enzymatic hydrolysis, various factors (NaOH concentration, solid content and pre-treatment time) that affect the pre-treatment process were investigated and evaluated based on the reducing sugar yield of the subsequent enzymatic hydrolysis. The enzymatic hydrolysis was based on the cellulase from Trichoderma reesi ATCC 26921, the optimum NaOH pre-treatment conditions were an NaOH concentration of 1.0% (w/w), a solid content of 5.0% (w/v) and a pre-treatment time of 60 min. Various parameters that affect the enzymatic hydrolysis of wheat straw, including the solid content, enzyme loading, pH and hydrolysis time, were investigated and optimized through a Box–Behnken design and response surface methodology. The predicted optimum conditions for enzymatic hydrolysis were a solid content of 8.0% (w/v), an enzyme loading of 35 FPU/g substrate, a temperature of 50 °C, a pH of 5.3 and a hydrolysis time of 96 h. The experimental result showed that the maximum reducing sugar yield was 60.73% (53.35% higher than the wheat straw without NaOH pre-treatment), which is in accordance with the predicted conditions.

Study on Optimization of Enzymatic Hydrolysis of Fish Bone Paste and Flavor Characteristics of Hydrolysates

Taking the ultra-fine fish bone paste prepared from cod bones as the object, the conditions of enzymatic hydrolysis of fish bone paste were optimized by using the single factor experiment and response surface design based on the degree of hydrolysis as an indicator. Meanwhile, the changes of flavor characteristics of fish bone paste and its hydrolysates were analyzed by employing electronic nose, electronic tongue, gas-phase ion migration chromatography(GC-IMS), and free amino acid determination. The results showed that the optimal enzymatic hydrolysis conditions were obtained at hydrolysis time of 5 hours, enzyme addition of 1.0%, material-liquid ratio of 1:1 by using flavor protease, yield 43.8% of hydrolysis degree. The electronic nose analysis results showed that the aromatic substances in the hydrolysate increased, while the content of undesirable compounds such as alkanes and ammonia decreased. The results of electronic tongue analysis showed that the acidity of the hydrolysate was increased, while the bitterness was reduced, and the overall taste was good and rich. There were 29 volatile compounds of hydrolysate and fish bone paste were detected by GC-IMS. In fish bone paste, the main volatile flavor substances were aldehydes and ketones, and the main flavor substances in the hydrolysate were ketones and esters. These types and contents of volatile flavor substances were significantly different. The hydrolysate increased the content of flavored amino acids and the hydrolysate had a good flavor.

A synergistic hydrothermal-deep eutectic solvent (DES) pretreatment for rapid fractionation and targeted valorization of hemicelluloses and cellulose from poplar wood

A synergistic pretreatment that realizing effective fractionation and targeted valorization can guarantee the implementability to future biorefinery scenario. In the present study, a stepwise approach using hydrothermal and deep eutectic solvents (DES) pretreatment was developed to preferentially dissociate hemicelluloses and further remove lignin from poplar, while retaining a cellulose-rich substrate that can be easily digested via enzymatic saccharification to obtain glucose. Results showed that the hydrothermal filtrate is mainly composed of xylooligosaccharide (XOS), monosaccharides, byproducts, and xylan-type hemicelluloses, which have homogenous structures and uniform molecular weights distribution as well as excellent antioxidant activity. Subsequent DES pretreatment further removed the lignin barriers, leading to a remarkable increase in the saccharification efficiency from 15.72% to 96.33% under optimum conditions for enzymatic hydrolysis. In short, the integrated pretreatment is effective for dissociating and chemical conversion of poplar wood, which was reasonable to promote the frontier of highly available biorefinery.

Novel peptides with α-glucosidase inhibitory activity from Changii Radix hydrolysates

The objective of this research was to investigate the α-glucosidase inhibitory activity of peptides released from Changii Radix protein under the optimal enzymatic hydrolysis conditions of trypsin, flavourzyme, compound proteinase, papain, alkaline proteinase, neutral protease, and in vitro simulated human gastrointestinal digestive conditions. The hydrolysate with the best activity was further purified by ultrafiltration, size-exclusion chromatography and semi-preparative high performance liquid chromatography. The peptides in the most active fraction were subsequently identified by nano-LC–MS/MS and screened for bioactivity. As a result, the GID-3-SG-2-RP-2 fraction (with smaller molecular weight and more hydrophobic) manifested the highest effect with an IC50 value of 32 μg/mL. Three peptides of this fraction were synthesized, SQHISTAGMEASGTSNMKF, KVIISAPSKDAPMF, and STFQQMW, and provided an IC50 of 1.051, 0.032, and 1.104 mg/mL, respectively. Thus, the results indicated that Changii Radix derived peptides may have potential functions in the prevention and management of type 2 diabetes.

Production and transepithelial transportation of casein-derived peptides and identification a novel antioxidant peptide LHSMK

In this study, casein derived peptides were hydrolyzed with trypsin and then transported across Caco-2 cell monolayers to choose intestinal absorbable antioxidant peptides. E/S ratio at 5.45 %, pH at 7.13 and temperature at 51.99 °C were found to be the optimal conditions for enzymatic hydrolysis of casein by trypsin by Response Surface Method. The fraction below 10 kDa purified by the hollow fiber ultrafiltration membrane was applied to Sephadex G-10 gel filtration chromatography, and the obtained components were transported across the Caco-2 cell monolayer. The transported peptides were detected on the basolateral side of the Caco-2 cell monolayer using ultra-performance liquid chromatography-tandem mass spectrometry. It was found that LHSMK was an unreported pentapeptide. The synthetic peptide LHSMK had free radical scavenging ability, and the antioxidant activity exhibited a dose-dependent increase. Furthermore, LHSMK can also regulate antioxidant enzymes activity in oxidative-damage cells. This study provides a method for the preparation of intestinal absorbable peptides, and provides basic research for the development of functional food market.