Composition Of Hydrolysates Research Articles

Effects of walnut kernel pellicle on the composition and properties of enzymatic hydrolysates of walnut meal by peptidomics and bioinformatics.

The purpose of this article is to investigate the effects of walnut (Juglans regia L.) kernel pellicle on the composition and properties of enzymatic hydrolysis products of walnut meal using peptidomics and bioinformatics. In this study, a total of 3423 peptide sequences were identified in peeled walnut protein hydrolysates (PWPH) and unpeeled walnut protein hydrolysates (UWPH). Due to the presence of the walnut kernel pellicle, the enzyme cleavage sites of alkaline proteases on walnut precursor proteins were altered, resulting in differences in the number and length of the peptides obtained. Principal component analysis indicates significant differences between PWPH and UWPH. Combined with bioinformatics analysis, it was shown that walnut kernel peeling improved the release of peptides, formed more bioactive peptides, reduced allergenicity, and improved water solubility. Seven peptides with acetylcholinesterase (AChE) inhibitory activity were identified, and the peptide Val-Gly-Ala-Pro-Phe-Asp-Gly-Ala (VGAPFDGA) has the strongest inhibitory activity with an IC50 of 0.38±0.01mg/mL. These results confirmed that walnut kernel peeling could greatly change the composition of the walnut protein hydrolysates, and seven novel peptides were reported that showed significant AChE inhibitory activity.

Integrated production of ethanol and 1,3-propanediol from food waste enzymatic hydrolysates in a biorefinery approach

The sustainable management of Urban Solid Waste (USW) is one of the most relevant global environmental challenges. Food waste (FW) constitutes a significant fraction of USW. One way to valorize FW is by producing biofuels and high-value-added chemicals. Therefore, this study aimed to produce 1,3-propanediol (1,3-PDO) from vinasse obtained after ethanolic fermentation of FW. Following the chemical characterization, the optimum composition of hydrolysate was generated from experimental design 23 + 3(0). Assay 7, with 69.12 g/L of glucose and 2.60 g/L of glycerol, was selected from the statistical analyses. Fermentation assays by Meyerozyma caribbica produced 26.88 g/L of ethanol and 6.36 g/L of glycerol. After ethanolic distillation, Lentilactobacillus diolivorans produced 1,3-propanediol with 74.28 % efficiency. This shows that the integrated production of ethanol and 1,3-PDO is a promising alternative for the biorefinery of FW.

Composition of Lignocellulose Hydrolysate in Different Biorefinery Strategies: Nutrients and Inhibitors

The hydrolysis and biotransformation of lignocellulose, i.e., biorefinery, can provide human beings with biofuels, bio-based chemicals, and materials, and is an important technology to solve the fossil energy crisis and promote global sustainable development. Biorefinery involves steps such as pretreatment, saccharification, and fermentation, and researchers have developed a variety of biorefinery strategies to optimize the process and reduce process costs in recent years. Lignocellulosic hydrolysates are platforms that connect the saccharification process and downstream fermentation. The hydrolysate composition is closely related to biomass raw materials, the pretreatment process, and the choice of biorefining strategies, and provides not only nutrients but also possible inhibitors for downstream fermentation. In this review, we summarized the effects of each stage of lignocellulosic biorefinery on nutrients and possible inhibitors, analyzed the huge differences in nutrient retention and inhibitor generation among various biorefinery strategies, and emphasized that all steps in lignocellulose biorefinery need to be considered comprehensively to achieve maximum nutrient retention and optimal control of inhibitors at low cost, to provide a reference for the development of biomass energy and chemicals.

Effect of bi-enzyme hydrolysis on the properties and composition of hydrolysates of Manchurian walnut dreg protein

Walnut dreg (WD) active peptides are an important source of dietary antioxidants; however, the products of conventional hydrolysis have limited industrial output owing to poor flavour and low bioactivity. To this end, in this study, we aimed to employ bvLAP, an aminopeptidase previously identified in our research, as well as commercially available Alcalase for bi-enzyme digestion. The flavour, antioxidant activity, and structures of products resulting from various digestion methods were compared. The results showed that the bi-enzyme digestion products had enhanced antioxidant activity, increased β-sheet content, and reduced bitterness intensity from 9.65 to 6.93. Moreover, bi-enzyme hydrolysates showed a more diverse amino acid composition containing 1640 peptides with distinct sequences. These results demonstrate that bi-enzyme hydrolysis could be a potential process for converting WD into functional food ingredients. Additionally, our results provide new concepts that can be applied in waste processing and high-value utilisation of WD.

Modulation of a Loop Region in the Substrate Binding Pocket Affects the Degree of Polymerization of Bacillus subtilis Chitosanase Products.

The hydrolytic products of chitosanase from Streptomyces avermitilis (SaCsn46A) were found to be aminoglucose and chitobiose, whereas those of chitosanase from Bacillus subtilis (BsCsn46A) were chitobiose and chitotriose. Therefore, the sequence alignment between SaCsn46A and BsCsn46A was conducted, revealing that the structure of BsCsn46A possesses an extra loop region (194N-200T) at the substrate binding pocket. To clarify the impact of this loop on hydrolytic properties, three mutants, SC, TJN, and TJA, were constructed. Eventually, the experimental results indicated that SC changed the ratio of chitobiose to chitotriose hydrolyzed by chitosanase from 1:1 into 2:3, while TJA resulted in a ratio of 15:7. This experiment combined molecular research to unveil a crucial loop within the substrate binding pocket of chitosanase. It also provides an effective strategy for mutagenesis and a foundation for altering hydrolysate composition and further applications in engineering chitosanase.

Non-animal protein hydrolysates from agro-industrial wastes: A prospect of alternative inputs for cultured meat

In light of the growing demand for alternative protein sources, laboratory-grown meat has been proposed as a potential solution to the challenges posed by conventional meat production. Cultured meat does not require animal slaughter and uses sustainable production methods, contributing to animal welfare, human health, and environmental sustainability. However, some challenges still need to be addressed in cultured meat production, such as the use of fetal bovine serum for medium supplementation. This ingredient has limited availability, increases production costs, and raises ethical concerns. This review explores the potential of non-animal protein hydrolysates derived from agro-industrial wastes as substitutes for critical components of fetal bovine serum in cultured meat production. Despite the lack of standardization of hydrolysate composition, the potential benefits of this alternative protein source may outweigh its disadvantages. Future research holds promise for increasing the accessibility of cultured meat.

Protein hydrolysate as a source of bioactive peptides in diabetic food products

Protein hydrolysates are a promising active component in specialized food products. Due to the presence of bioactive peptides with enzyme inhibiting activity in their composition, protein hydrolysates exert different physiological effects. The aim of the research was to establish the potential hypoglycemic activity of the protein hydrolysate of broiler chicken stomachs in whey and to develop a beverage on its basis for patients with diabetes mellitus. Protein hydrolysates obtained from microbial fermentation of muscular stomachs of broiler chickens in whey were assessed by UHPLC coupled to mass-spectrometry. Using the BioPEp database, the molecular weight distribution of peptides in the hydrolysates was determined. In addition, bioactive peptides showing the inhibitory activity toward enzymes dipeptidyl peptidase‑4 and amylase were revealed. Based on the obtained hydrolysates, recipes of beverages for patients with diabetes mellitus were modeled in the Excel software program with regard to the recommendations for nutrition of this population group. According to the obtained recipes, beverages were produced in the laboratory conditions and their taste was assessed. An effect of the beverage on glucose reduction in the blood of laboratory animals was assessed upon single peroral administration with a dose of 1.3 ml before the main meal. The results of the investigation of the peptide composition of protein hydrolysates showed the presence of bioactive peptides (SY, VW, SW) in them with the inhibitory activity toward enzymes dipeptidyl peptidase‑4 and amylase confirmed by the BioPep database. The highest amount of the above indicated bioactive peptides (37.1 mg/100 g hydrolysate) was in the protein hydrolysate produced by fermentation of raw materials with propionic acid bacteria. The beverage produced on the basis of the protein hydrolysate and lemon juice was more acceptable for consumers in terms of taste than the beverage with the taste of cacao. The results of the in vivo trials showed that the beverage based on the protein hydrolysate was effective in reducing sugar in blood of both healthy rats and rats with diabetes mellitus. The blood sugar level in the healthy animals of the G1 group (without using the beverage) increased by 151% relative to the initial values, which was a higher value compared to the G2 group (with the use of the beverage), where an increase was 87%. A more significant growth in the blood sugar level (61% relative to the initial values) was also observed in the ill animals in the G3 group (without using the beverage), while this increase was 46% in the G4 group (with the use of the beverage). Thus, the results of the study show the prospects of using the protein hydrolysates from broiler chicken stomachs in whey as an active hypoglycemic component in beverages for patients with diabetes mellitus. However, more profound research is necessary including studies on the representative group of patients.

Effect of hydrolysis degree with Alcalase on antioxidant and antigenic properties of whey and colostrum protein hydrolysates

Enzymatic hydrolysis of dairy proteins is aimed to increase their nutritional value and reduce their allergenic properties. It is relevant to study the effects of enzymatic hydrolysis with highly active endopeptidase alсalase on the bioactivity of the whey and colostrum protein components. Dairy hydrolysates with various degree of Alcalase treatment were obtained in the present study. A comparative analysis of cleaved dairy proteins and their peptide fractions with a 5 and 10 kDa cutoff was performed. The protein and peptide composition of hydrolysates was determined using various techniques. The bioactivity of hydrolyzed whey and colostrum samples were estimated. The antioxidant activity of hydrolyzed and fractionated (5 kDa cutoff) whey samples increased by 5.91/6.62-fold and that of colostrum by 14.22/7.24-fold in fluorimetric/spectrophotometric assay, respectively. A significant decrease in the antigenic potential of milk peptides was observed with increasing degree of hydrolysis and subsequent ultrafiltration. Thus, bioactive dairy protein hydrolysates were obtained, which are applicable as a component of specialized foods with proven hypoallergenic properties and high antioxidant activity.

Protein hydrolysates from Mytilus edulis L. mussel: Physicochemical and antioxidant properties

Meat of marine molluscs is a promising protein-containing raw material for obtaining protein hydrolysate and biologically active peptides. The purpose of this research was to study the physicochemical properties and antioxidant properties of protein hydrolysate obtained from the mussel Mytilus edulis L. The hydrolysates were obtained by acidic hydrolysis and enzymatic hydrolysis using protozyme as an enzyme. Gel permeation chromatography was used as experimental method in this study to characterise the molecular weight distribution of the protein hydrolysates. Chemical analysis was used for determine the composition and antioxidant activity of hydrolysates. It is shown that the content of low-molecular fractions - biologically active peptides with a molecular weight of less than 5 kDa is almost three times higher in the enzymatic hydrolysate compared to the acidic one. Accordingly, the enzymatic hydrolysate demonstrates high antioxidant activity, which determines its prospects as a component for functional food products and biomedical drugs.

Protein Hydrolysates from Fishery Processing By-Products: Production, Characteristics, Food Applications, and Challenges.

Fish processing by-products such as frames, trimmings, and viscera of commercial fish species are rich in proteins. Thus, they could potentially be an economical source of proteins that may be used to obtain bioactive peptides and functional protein hydrolysates for the food and nutraceutical industries. The structure, composition, and biological activities of peptides and hydrolysates depend on the freshness and the actual composition of the material. Peptides isolated from fishery by-products showed antioxidant activity. Changes in hydrolysis parameters changed the sequence and properties of the peptides and determined their physiological functions. The optimization of the value of such peptides and the production costs must be considered for each particular source of marine by-products and for their specific food applications. This review will discuss the functional properties of fishery by-products prepared using hydrolysis and their potential food applications. It also reviews the structure-activity relationships of the antioxidant activity of peptides as well as challenges to the use of fishery by-products for protein hydrolysate production.

The Effects of Enzymes, Species, and Storage of Raw Material on Physicochemical Properties of Protein Hydrolysates from Whitefish Heads.

The rest raw materials of whitefish have great potential for increased utilisation and value creation. Whitefish heads have a high protein content and should be considered a healthy protein source for the growing population's demands for sustainable protein. In this study, the heads of four different species of whitefish were processed via enzymatic hydrolysis, namely cod (Gadus morhua), cusk (Brosme bromse), haddock (Melanogrammus aeglefinus), and saithe (Pollachius virens), using three commercially available enzymes. Trials were conducted after 0, 3, and 6 months of the frozen storage of heads. A proximate analysis, molecular weight distribution, and protein solubility were evaluated for each of the products. The results show that, although the enzymatic hydrolysis of rest raw materials from different species of whitefish yielded products of slightly different characteristics, this process is viable for the production of high-quality protein from cod, cusk, haddock, and saithe heads. Six months of frozen storage of heads had a minimal effect on the yield and proximate composition of hydrolysates.

Investigation of the biopotential of products of hydrolysis of waste from cutting the white-legged shrimp Penaeus vannamei

The relevance of studying the biopotential of shrimp waste and their hydrolysates is due to the need for complex processing of secondary raw materials of aquatic organisms to obtain useful products. At the fish processing plant Vichyunai-Rus LLC when manufacturing food products from white-legged shrimp, up to 60 % of the mass of waste (cephalothorax, shell) remains. This raw material contains valuable organic components, but is not processed. The paper proposes its complex processing with the production of hydrolysates in two ways – high-temperature and enzymatic. During thermohydrolysis in the aquatic environment, three fractions are formed from shrimp waste (fatty, water-soluble and water-insoluble). After separation, the water-soluble fraction was sublimated, and the water-insoluble fraction was dried by convection. The fat fraction was further purified by washing in warm water. The general chemical composition of shrimp waste and hydrolysis products has been studied. It has been shown that water-soluble hydrolysates are a good source of protein components (66.6–71.6 %). In comparative studies of the amino acid composition of water-soluble hydrolysates, the presence of all essential amino acids is established with minor differences between the samples. Both hydrolysates are dominated by alanine, arginine, glycine, isoleucine, lysine, aspartic acid, tyrosine, valine (3.3–6.4 g/100 g of protein). In terms of formol-titratable nitrogen in fermentolisates, alkalase ferments shrimp waste more actively than collagenase. The fatty acid composition of lipids isolated by thermohydrolysis from shrimp waste has been analyzed. Shrimp oil is characterized by a high content of polyunsaturated fatty acids (44.7 %) with a relatively low content of omega-3 family acids (10.7 %) and a high content of omega-6 (33.9 %) at a ratio of 1 : 3.2 (close to the physiologically recommended). The organoleptic characteristics of water-soluble and water-insoluble shrimp hydrolysates have been studied. Due to the content of valuable biologically active components in hydrolyzates, their use as food and feed additives – sources of active peptides, high-molecular proteins, minerals and chitinous components – is recommended.

Nutritional composition, health benefits and bio-active compounds of chickpea (Cicer arietinum L.).

Chickpea (Cicer arietinum L.), an annual plant of the family Fabaceae is mainly grown in semiarid and temperate regions. Among pulses, cultivated worldwide chickpeas are considered an inexpensive and rich source of protein. Chickpea is a good source of protein and carbohydrate, fiber, and important source of essential minerals and vitamins. The quality of protein is better among other pulses. Consumption of chickpeas is related to beneficial health outcomes. Dietary peptides from the protein of chickpeas gaining more attention. Peptides can be obtained through acid, alkali, and enzymatic hydrolysis. Among all these, enzymatic hydrolysis is considered safe. Various enzymes are used for the production of peptides, i.e., flavorzyme, chymotrypsin, pepsin, alcalase, papain, and trypsin either alone or in combinations. Chickpea hydrolysate and peptides have various bioactivity including angiotensin 1-converting enzyme inhibition, digestive diseases, hypocholesterolemic, CVD, antioxidant activity, type 2 diabetes, anti-inflammatory, antimicrobial, and anticarcinogenic activity. This review summarizes the nutritional composition and bioactivity of hydrolysate and peptides obtained from chickpea protein. The literature shows that chickpea peptides and hydrolysate have various functional activities. But due to the limited research and technology, the sequences of peptides are unknown, due to which it is difficult to conduct the mechanism studies that how these peptides interact. Therefore, emphasis must be given to the optimization of the production of chickpea bioactive peptides, in vivo studies of chickpea bioactivity, and conducting human study trials to check the bioactivity of these peptides and hydrolysate.

Calanus finmarchicus as a novel source of health-promoting bioactive peptides: Enzymatic protein hydrolysis, characterization, and in vitro bioactivity

Calanus finmarchicus is a crustacean currently used as a source of marine lipid. The lipids are extracted by enzymatic protein hydrolysis, while the remaining peptide fraction is regarded as a byproduct. In the present work, ten different commercial proteases and endogenous C. finmarchicus proteases were used to produce a set of 63 protein hydrolysates. Protease concentration and hydrolysis time were varied. Hydrolysates were characterized using size-exclusion chromatography and 1H nuclear magnetic resonance spectroscopy. Addition of commercial proteases had unremarkable effect on the yield and molecular weight distribution. This was attributed to the strong impact of endogenous enzymes dominating the hydrolysis process. However, multivariate classification based on 1H NMR spectra revealed subtle variations in composition of hydrolysates produced using different enzymes. The hydrolysates were further evaluated for DPP-IV inhibition and antioxidant activity. The hydrolysates showed significantly higher bioactivity than the unhydrolyzed control. A representative hydrolysate (CaFi55) was fractionated using semipreparative size-exclusion chromatography. A fraction consisting of short peptides with an average chain length of five amino acids (F2), was identified as a major contributor to the DPP-IV inhibitory activity (IC50 = 0.70 ± 0.07 mg/mL).

Health-promoting activities of Moringa oleifera Lam. seeds protein hydrolysates and its ultra-filtered peptide fractions

Moringa oleifera (MO) seeds treated as a new resource food rich in protein (37.48%), but, little know the bioactive benefits and peptide composition of protein hydrolysates. In this study, ultrafiltrated peptide fractions of MO seed proteins hydrolysates (MOSPH) were analyzed for their antioxidant, anti-hypertension and sleep-promoting activities. LC-MS/MS was used to identify the peptide composition of the molecular weight (MW) <3 kDa fraction. The results showed that MOSPH and the lower MW peptides fraction, especially the MW <3 kDa fraction, exhibited higher free radical scavenging and reducing capacity, in vitro angiotensin I-converting enzyme (ACE) inhibitory capacity and vasodilatory and anti-hypertensive effects on spontaneously hypertensive rats (SHR) after the development of hypertension, sleep-promoting and anticonvulsant effects in male ICR mice. Furthermore, four potential bioactive peptides (pepsite score > 0.3) from MW <3 kDa fraction of MOSPH were explored via in silico approach, of which a novel dual-function peptide IWHHTFYNELR (IR-11) exhibited higher ACE inhibitory activity (IC50 = 1.057 mg/mL) and DPPH•-clearing activity (IC50 = 0.404 mg/mL). Molecular docking indicates that IR-11 interacted with ACE and DPPH• through hydrogen bonding and hydrophobic forces. Our results suggested that MO seed proteins peptides can be a potent functional agents for developing new functional food.

Transforming Spent Coffee Grounds' Hydrolysates with Yeast Lachancea thermotolerans and Lactic Acid Bacterium Lactiplantibacillus plantarum to Develop Potential Novel Alcoholic Beverages.

In the present work, the modification of spent coffee grounds (SCG) hydrolysate composition by mixed cultures of a non-Saccharomyces yeast, Lachancea thermotolerans, and a lactic acid bacterium, Lactiplantibacillus plantarum, as well as their interactions, were evaluated. It was found that L. plantarum inhibited the growth and survival of L. thermotolerans as compared with that in the yeast alone. On the other hand, the growth and survival of L. plantarum was slowed in sequential fermentation, but not in co-culture. Compared with co-culture, higher ethanol content, less residual sugars, and less acetic and succinic acids were found in sequential fermentation. In addition, lower amounts of caffeine and phenolic acids (e.g., ferulic, caffeic, and p-coumaric acids) were obtained in mixed (co- and sequential) cultures with corresponding levels of volatile phenols relative to the yeast monoculture. Moreover, co-culturing resulted in the highest contents of total alcohols (ethanol excluded) and total esters. Therefore, mixed culturing of L. plantarum and L. thermotolerans presented positive effects on the chemical constituents of fermented SCG hydrolysates, which might be a new alternative approach to valorizing the SCG into novel alcoholic drinks with different ethanol and flavor constituents.

Molecular Docking and Site-Directed Mutagenesis of GH49 Family Dextranase for the Preparation of High-Degree Polymerization Isomaltooligosaccharide.

The high-degree polymerization of isomaltooligosaccharide (IMO) not only effectively promotes the growth and reproduction of Bifidobacterium in the human body but also renders it resistant to rapid degradation by gastric acid and can stimulate insulin secretion. In this study, we chose the engineered strain expressed dextranase (PsDex1711) as the research model and used the AutoDock vina molecular docking technique to dock IMO4, IMO5, and IMO6 with it to obtain mutation sites, and then studied the potential effect of key amino acids in this enzyme on its hydrolysate composition and enzymatic properties by site-directed mutagenesis method. It was found that the yield of IMO4 increased significantly to 62.32% by the mutant enzyme H373A. Saturation mutation depicted that the yield of IMO4 increased to 69.81% by the mutant enzyme H373R, and its neighboring site S374R IMO4 yield was augmented to 64.31%. Analysis of the enzymatic properties of the mutant enzyme revealed that the optimum temperature of H373R decreased from 30 °C to 20 °C, and more than 70% of the enzyme activity was maintained under alkaline conditions. The double-site saturation mutation results showed that the mutant enzyme H373R/N445Y IMO4 yield increased to 68.57%. The results suggest that the 373 sites with basic non-polar amino acids, such as arginine and histidine, affect the catalytic properties of the enzyme. The findings provide an important theoretical basis for the future marketable production of IMO4 and analysis of the structure of dextranase.

Production of Biomodified Bleached Kraft Pulp by Catalytic Conversion Using Penicillium verruculosum Enzymes: Composition, Properties, Structure, and Application

The global development of the bioeconomy is impossible without technologies for comprehensive processing of plant renewable resources. The use of proven pretreatment technologies raises the possibility of the industrial implementation of the enzymatic conversion of polysaccharides from lignocellulose considering the process’s complexity. For instance, a well-tuned kraft pulping produces a substrate easily degraded by cellulases and hemicelulases. Enzymatic hydrolysis of bleached hardwood kraft pulp was carried out using an enzyme complex of endoglucanases, cellobiohydrolases, β-glucosidases, and xylanases produced by recombinant strains of Penicillium verruculosum at a 10 FPU/g mixture rate and a 10% substrate concentration. As a result of biocatalysis, the following products were obtained: sugar solution, mainly glucose, xylobiose, xylose, as well as other minor reducing sugars; a modified complex based on cellulose and xylan. The composition of the biomodified kraft pulp was determined by HPLC. The method for determining the crystallinity on an X-ray diffractometer was used to characterize the properties. The article shows the possibility of producing biomodified cellulose cryogels by amorphization with concentrated 85% H3PO4 followed by precipitation with water and supercritical drying. The analysis of the enzymatic hydrolysate composition revealed the predominance of glucose (55–67%) among the reducing sugars with a maximum content in the solution up to 6% after 72 h. The properties and structure of the modified kraft pulp were shown to change during biocatalysis; in particular, the crystallinity increased by 5% after 3 h of enzymatic hydrolysis. We obtained cryogels based on the initial and biomodified kraft pulp with conversion rates of 35, 50, and 70%. The properties of these cryogels are not inferior to those of cryogels based on industrial microcrystalline cellulose, as confirmed by the specific surface area, degree of swelling, porosity, and SEM images. Thus, kraft pulp enzymatic hydrolysis offers prospects not only for producing sugar-rich hydrolysates for microbiological synthesis, but also cellulose powders and cryogels with specified properties.

Integrated rice bran processing by supercritical CO2 extraction and subcritical water hydrolysis to obtain oil, fermentable sugars, and platform chemicals

Rice bran is rich in oil and constituted by a lignocellulosic structure, which is a potential raw material for its processing into several products. Therefore, the oil yield and composition of oil obtained from supercritical CO2 extraction were investigated to select a condition for using defatted rice bran for processing by subcritical water hydrolysis. The effects of temperature (230 °C and 260 °C) and solvent-to-feed mass ratio (50 and 100 g water/g defatted rice bran) on fermentable sugar yield (YFS) and hydrolysate composition were evaluated and discussed. Supercritical CO2 extraction at 40 °C and 20 MPa provided an oil yield of 17.19 wt% at 120 min. For defatted rice bran, the highest YFS (6.53 g sugars/100 g defatted rice bran) was obtained at 260 °C and 100 g water/g defatted rice bran.

Production of dehairing protease by Bacillus cereus VITSN04: a model cradle-to-cradle approach for sustainable greener production of leathers

ABSTRACT Despite several attempts over decades, process scalability and sustainability remains a challenge to produce an environmental-friendly enzyme to gain industrial attention. In the present study, microbial degradation of chrome shavings (chromium-collagen leather waste) and the resulting collagen hydrolysate for producing the dehairing protease by Bacillus cereus VITSN04 were investigated in a lab-scale fermentor. Scale-up degradation of shavings resulted in higher recovery of collagen hydrolysate (76%) within 72 h compared to shake flasks (68% in 120 h). Earlier achieved medium composition of collagen hydrolysate (12 g L-1) and molasses (15 g L-1) appeared to induce amylase in high rate, despite maximal production of protease (203.8±0.18 U mL-1), which was analysed by ANS fluorescence spectroscopy. Optimization of the media containing collagen hydrolysate (12 g L-1) and molasses (5 g L-1) was effective in producing protease (170.6±0.1 U mL-1) and reduce the co-synthesis of amylase (48.2±0.09 U mL-1). The controlled fermentation process, by feeding molasses during exponential growth phase had enhanced dehairing protease production (∼2.96 fold). The produced protease then partitioned through biphasic system showed significant dehairing of goat skins in pilot scale. Thus, scalability of the process to produce dehairing enzymes using waste, generated at the site of its use offers hope for sustainable greener production of leathers.