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A novel metalloprotease from banana peel and its biochemical characterization
Abstract
Protein cleaving enzymes, called proteases are one of the most commercially used enzymes
possessing extensive applications in various industries. The present study was focused on
screening, extraction, purification and characterization of protease from banana peel. Twelve
different varieties of banana peels were screened for the protease activity. The maximum
activity of the isolated enzyme was 230.4 CDU/mg from Nendran banana. The crude enzyme
was purified up to 8.1 fold with the yield of 61.34%. The molecular weight of the purified
fraction was found to be 40 kDa. The optimum conditions for maximum protease activity were
achieved at 30°C and pH 7. The Lineweaver-Burk plot exhibited the kinetic parameters of the
enzyme such as Vmax and Km as 588.4 CDU/mg and 15.2 mg/mL respectively. The influence
of different inhibitors, metal ions, organic solvents, detergents, oxidising agents and salinity
were examined. The collagenolytic activity was tested with purified type I collagen as a
substrate. The enzyme was tested for cell cytotoxicity, detection of apoptosis using fluorescent
dyes and antitumor activity. The results demonstrated that protease from banana peel was of
high significance and potential for usage in therapeutic for breaking down collagen / peptide
bonds.
Keywords: Banana peel, Protease, Collagenase activity, MTT
... Gallic acid extracted from agricultural waste with the help of this method is a good by-product to be used as a food additive (Curiel et al. 2010). Gurumallesh et al. (2019) screened, extracted, purified, and characterized protease enzymes from banana peels. ...
... The unwanted fragments are separated during banana processing operations and can be further used to extract bioactive components (Mathew and Negi 2017;Vu et al. 2018). Gurumallesh et al. (2019) screened, extracted, purified, and characterized protease enzymes from banana peels. Twelve various species of banana peels were screened for protease enzyme activity. ...
Vegetables and fruits have been considered as one of the most consumed staple foods in society as these possess many health-promoting factors. The enhanced day-today demand has fastened their production and processing rate, which generates a plethora of significant fresh and processed vegetable and fruit wastes. Various vegetable and fruit processing practices produce around 20-30% of waste by-products constituted of skin, pomace, seeds, etc. These waste materials are a rich source of many functional bioactive compounds such as polyphenols, vitamins , fibers, enzymes, oils, etc. These phenolic and bioactive compounds can be recovered from vegetable and fruit wastes through several extraction methods. Further, these extracted valuable materials have been valorized into value-added products especially tailored for many pharmaceuticals, health, and food applications. The present chapter focuses on types of vegetable and fruit waste, phenolic and bioactive compounds present in this waste, various strategies from the extraction of these valuable materials, and their possible conversion into functional value-added products.
... Gallic acid extracted from agricultural waste with the help of this method is a good by-product to be used as a food additive (Curiel et al. 2010). Gurumallesh et al. (2019) screened, extracted, purified, and characterized protease enzymes from banana peels. ...
... The unwanted fragments are separated during banana processing operations and can be further used to extract bioactive components (Mathew and Negi 2017;Vu et al. 2018). Gurumallesh et al. (2019) screened, extracted, purified, and characterized protease enzymes from banana peels. Twelve various species of banana peels were screened for protease enzyme activity. ...
Reduction of food waste by extensively using raw sources builds good impact on the climate, environment, and food security. Fruits and vegetables are the commodities of horticultural crops that are commonly used. Thus, the waste generated during processing can be proclaimed as a valuable by-product if extensive technological interventions are used to improve the value of successive goods to balance the cost of processing. Fruit and vegetable wastes have great potential for bioconversion into valuable products with biotechnological, biocontrol, bioenergy, and industrial applications. These wastes could serve as a source of edible coating, fat mimetics, fortified probiotics, natural pigment, and volatile compounds in the food industry. Additionally, these wastes can also be used for synthesizing green carbon dots, biochar, and biosorbents that have environmental applications. The utilization of these fruit and vegetable wastes for generating valuable products is a foundation step towards sustainable development. The current chapter discusses the valuable products that can be obtained by bioconversion of waste of fruits and vegetables by enlisting their application in both food and environmental sectors.
... Gallic acid extracted from agricultural waste with the help of this method is a good by-product to be used as a food additive (Curiel et al. 2010). Gurumallesh et al. (2019) screened, extracted, purified, and characterized protease enzymes from banana peels. ...
... The unwanted fragments are separated during banana processing operations and can be further used to extract bioactive components (Mathew and Negi 2017;Vu et al. 2018). Gurumallesh et al. (2019) screened, extracted, purified, and characterized protease enzymes from banana peels. Twelve various species of banana peels were screened for protease enzyme activity. ...
Agricultural residues, including fruit and vegetable residues, which usually contain starch or cellulose, are used for bioconversion into value-added products. Bioconversion provides several advantages, such as high value considering their material and energy recovery, decrease in landfill areas, lower cost of the technology, and the farmers’ income. Bioprocessing is of great importance for horticultural waste management to satisfy the needs of developing countries. Fermentation (solid-state and submerged) is considered an important tool to increase the nutritional value (providing more proteins, amino acids, and other biomolecules) of SCP and aquafeed meal ingredients. This chapter investigates the role of fruit and vegetable wastes in the production of single-cell protein (SCP) and aquafeed meal.
... Bananas are not only sold as fresh fruits but are also utilised in various processed products [5,6]. However, the amount of biomass generated during banana preparation is enormous; in fact, the peel accounts for 35-40% of the total weight [7]. In Thailand, 'Nam wa mali-ong' bananas are an important variety due to their desirable ...
Banana peel (BP) is the primary by-product generated during banana processing which causes numerous environmental issues. This study examines the physical attributes, proximate analysis, glycoarray profiling, antioxidant abilities, and prebiotic activity of BP. The analysis demonstrated that carbohydrates constituted the primary components of BP and the glycoarray profiling indicated that BP contains multiple pectin and hemicellulose structures. BP also contained phenolic compounds, including (+)-catechin and gallic acid, flavonoid compounds, and antioxidant activities. BP demonstrated prebiotic effects by promoting the proliferation of advantageous gut bacteria while inhibiting the growth of harmful bacteria. The prebiotic index scores demonstrated that BP exhibited a greater capacity to promote the growth of beneficial bacteria in comparison to regular sugar. The study demonstrated the potential of the BP as a valuable source of dietary fibre, bioactive compounds, and prebiotics. These components have beneficial characteristics and can be utilised in the production of food, feed additives, and functional food.
... Banana plant wastes have been used in numerous studies as novel culinary additives, alternative energy sources, or critical components for water and sewage treatment. Banana peels have also been used by researchers to make antidepressant candies (Gurumallesh et al., 2019) and milk beverages (Batista et al., 2017). Banana plants can be used in the production of long-lasting battery cells as an environmentally friendly option , as a component of bioethanol, or as an additive to improve the production of biogas (Guerrero et al., 2018). ...
Hydrogen sulfide (H2S) gaseous pollutants are naturally generated from natural gas streams, organic waste degradation, and biogas production. Removal of the H2S pollutants is very important due to their harmful and odorous gas. In this study, the removal of this gas was carried out with a natural biosorbent that is abundant in tropical countries, namely banana leaves. This study aimed to reduce H2S gas through the adsorption process using different types of banana leaves, specifically Kepok Putih, Raja Nangka, and Tonto. This study used adsorption tubes and measurements of H2S with a spectrophotometer and tested the characteristics of the leaves with SEM–EDX. The results showed that all tested banana leaves here performed removal of H2S gas, albeit at different levels. The H2S gas adsorption efficiency values for the Kepok Putih, Raja Nangka, and Tonto leaves were estimated to be 45.66%, 72.93%, and 59.08%, respectively. The best adsorbents were Raja Nangka banana leaves, which had an 11.0480 mg/g adsorption capacity. Based on this study’s results, it seems that leaf stomatal size determines adsorption capacity compared to moisture and surface area of the banana leaf. The abundance of this biosorbent in rural areas and ease of preparation make it feasible to be developed as a low-cost H2S biosorbent.
... The extract from banana peel also exhibits strong antioxidant, anti-fungal, and anti-bacterial activity. According to Gurumallesh et al., (2019), banana peel possesses a novel metalloprotease which exerts anti-carcinogenic activity through the break down of collagen peptide bonds. Lemon peel contains flavones (apigenin-glucoside, diosmetin-glucoside), limonoids (ichangin), and flabanones (eriocitrin and hesperidin). ...
Garden herbs have medicinal value and were used traditionally to promote well-being. It also means that herbs were in fact constituents of native diets. However, modern societies have neglected the benefits of herbs and rely on other food sources for nutrition. This study has examined a total of eight herbs; Amomum subulatum, Calendula officinalis, Cymbopogon flexuosus, Eryngium foetidum, Lantana indica,Ocimum bacilicum, Ocimum gratissimum and Salvia mirzayanii from a vast collection of 19 published works by using systematic review. The findings inform that herbs could be prepared in both, dry-form and extracts before being used in various applications. However, it was the essential oils of these herbs that divided them into categories such as safe for consumption and safe for industrial applications. The compounds of these essential oils were later evaluated for either medicinal, anti- microbial or for use in insecticide. The basis of most compounds was aromatic which means that all herbs are rich with phenols, oils and scent. These constituents enabled the herbs to have antioxidant properties. Through a scrutiny, only A. subulatum, C. officinalis, E. foetidum and O. bacilicum could be promoted for supplementary use because these herbs were extensively studied for oral intake. The findings of this work are crucial for the bio-extractive industry because not all herbs although used traditionally, could be processed into supplements to promote well- being.
... The metalloprotease from banana peel was extracted and the isolation procedure and its biochemical characterization were explained 11 . The varieties of banana used for screening protease activity were Nendran, Nadan, Kadhali, Poovan, Yelaki, Rastali, Karpooravalli, Pacha nadan, Virupaskshi, Red banana, Morris (Yellow), and Morris (green), which are local names in Tamil language. ...
Proteases are a group of enzymes that cleaves on peptide bonds specifically. Proteolytic enzymes are of global importance because of their widespread applications in therapeutics and commercial purposes. Despite existing proteases, researchers are focusing in finding some cost effective protease source to meet out the demand in the enzyme market. Amongst the various sources available, nendran banana peel had been identified as a potential source of metalloprotease. The isolated biomolecules could be immobilized into various physical forms and has extensive potential in diverse fields. The characterization of the isolated enzyme was analyzed using particle size analyzer, TEM, TGA, DSC, MALDI-TOF and proton NMR. Based on the physical parameters techniques the right vehicle could be finalized. The average particle size of the molecules was found to be 2110 d.nm with a poly dispersity index of 0.299 and the intercept is 0.860. The zeta potential was found to be -8.93 mV with a zeta deviation of 15.6 mV and conductivity of 4.03 (mS/cm). From the results of TEM it is well clear that there are no signs of agglomeration among the particles. With the help of TGA and DSC studies it is well understood that the proteolytic enzymes have the utmost thermal stability. The results suggest that the enzyme could add a new insight on protease research.
... In most SSF processes, the carbon and energy source is a natural raw resource. SSF can also use an inert substance as a solid matrix; however this involves adding extra nutrients and a carbon source to the nutrition solution [7,8]. However, the solid substrate (matrix) needs to have enough moisture. ...
The Actinomycetes are gram-positive, unicellular bacteria that grow in a typical filamentous manner on solid substrates and may synthesis a wide range of secondary metabolites, including enzymes. Proteases are universally present enzymes that catalyze hydrolytic processes that break down protein molecules into peptides and amino acids. The protease enzyme is used in food and detergent industries. The goal of the current investigation is to identify and isolate a prospective isolate of Actinomycetes capable of solid state fermentation for the protease enzyme production. The soil samples yielded a total of 30 isolates. On starch casein nutrition agar medium, B5 isolate was observed to be a potential strain for production of protease enzyme. The optimum conditions for the maximum protease enzyme production were observed to be at 30°C with a pH of 7 when incubated for 5 days with a 6th day old culture and 2mL of the inoculum volume for protease with substrate concentration (pomegranate and banana peel powder of each 2.5 g) of 5 g. The addition of Carbon source starch and Nitrogen source casein to the production medium has enhanced the enzyme activity to 442.8 U/gds.
... Fat and carbohydrate are good sources of energy for livestock and humans. Crude fibre content in diets have been reported to aid digestive processes; promoting health in livestock and humans (Abubakar et al., 2016) and these findings are in line with previous reports indicating the bioactivity and application of peels of various plants (Abbas et al., 2018;Abdel Aziz et al., 2018;Ahad et al., 2018;Derakhshan et al., 2018;Garrafa-Galvez et al., 2019;Gurumallesh et al., 2019;Hu et al., 2019;Jridi et al., 2019;Kharchoufi et al., 2018;Kokila et al., 2016;Lakkab et al., 2019;Liu et al., 2017;Rodsamran and Sothornvit, 2019;Xu et al., 2019). ...
This study investigated the nutraceutical potential of ripe and unripe plantain
fruit peels which are commonly discarded as food wastes. Proximate and mineral
analyses of the samples were performed as per the standard methods of the
Association of Official Analytical Chemists. Preliminary phytochemical screening
of aqueous, acetone and methanol extracts of the peels was also carried out in
accordance to standard methods. From the results of the study, acetone extract of
the unripe peel showed the presence of eight phytochemicals while its ripe peel
showed the presence of four. Aqueous and ethanol extracts of both peels showed
the presence of same phytochemicals i.e., terpenoids, cardiac glycosides, phenols,
flavonoids, alkaloids, reducing sugars and saponins. Meanwhile, tannins was
absent in all three solvent extracts of both peels. Fat, ash, crude fibre and
carbohydrate contents of the unripe peel were higher than those of the ripe.
However, moisture and protein contents of the ripe peel were significantly higher
(P < 0.05) than those of the unripe. Of all the nine essential minerals assayed (K,
Na, Mg, Ca, P, Fe, Zn, Mn, Cu), concentrations of all except calcium were
significantly higher (P < 0.05) in the unripe peel than those of the ripe peel.
Notably, none of the heavy metals (Co, Cr, Cd, Pb, Ni) assayed was detected in
both samples. This study concludes that ripe and unripe plantain fruit peels could
serve as promising sources of nutrients and bioactive compounds essential for
the health of both livestock and humans
... Vu, Scarlett, and Vuong (2019) also found that different ripening stages substantially impact the antioxidant potential of banana peel extract, with antioxidant activity increasing in ripe bananas and decreasing in overripe fruits. A new metalloprotease isolated from banana peel has shown the potential to be employed in cancer treatment (its mechanism involves breaking down of collagen peptide bonds) (Gurumallesh, Ramakrishnan, and Dhurai 2019). ...
Nanotechnology is a rapidly growing field with profound applications in different domains, particularly in food science and technology. Nanoparticles (NPs) synthesis, an integral part of nanotechnology-based applications, is broadly classified into chemical, physical and biosynthesis methods. Chemically sensitive and energy-intensive procedures employed for NPs synthesis are some of the limits of traditional chemical approaches. Recent research has focused on developing easy, non-toxic, cost-effective, and environment-friendly NPs synthesis during the last decade. Biosynthesis approaches have been developed to achieve this goal as it is a viable alternative to existing chemical techniques for the synthesis of metallic nanomaterials. Fruit peels contain abundant bioactive compounds including phenols, flavonoids, tannins, triterpenoids, steroids, glycosides, carotenoids, anthocyanins, ellagitannins, vitamin C, and essential oils with substantial health benefits, anti-bacterial and antioxidant properties, generally discarded as byproduct or waste by the fruit processing industry. NPs synthesised using bioactive compounds from fruit peel has futuristic applications for an unrealized market potential for nutraceutical and pharmaceutical delivery. Numerous studies have been conducted for the biosynthesis of metallic NPs such as silver (AgNPs), gold (AuNPs), zinc oxide, iron, copper, palladium and titanium using fruit peel extract, and their synthesis mechanism have been reported in the present review. Additionally, NPs synthesis methods and applications of fruit peel NPs have been discussed.
... Researchers have also used banana peels as antidepressant candy [2] or as an ingredient for milk drinks [3]. The banana plant can also be used as an environmentally friendly alternative for long-lasting battery cells [4], as a raw material for producing bioethanol, or as a complementary material to increase biogas production [5]. ...
Bananas have the highest production rate among fruits in Indonesia, which leads to the generation of a significant amount of banana fruit solid waste. In this study, we assessed the potential use of banana waste to remove hydrogen sulfide (H2S) gas. In particular, the purpose of this study was to analyze the efficiency of banana waste as an adsorbent for H2S gas. We tested the stems, leaves, and peels of banana plants as H2S gas adsorbents with varying contact times. To obtain a microscopic view of the adsorbents before and after the experiment, we conducted measurements using scanning electron microscopy with dispersive X-ray spectroscopy. The banana leaves, stems, and peels were found to have H2S gas absorption efficiency values of 76.52%, 51.83%, and 6.44%, respectively. Based on the experiment, the leaves of the banana plant appear to be the best adsorbents, with an adsorption capacity of 1.67 mg/g. The results also revealed that there was a change in the fiber and stomata appearance of the banana leaves after the adsorption process. Overall, this research indicates that banana leaves have the potential to be used as effective H2S adsorbents.
... Furthermore, among several kinds of pectin-containing by-products, banana peel was observed as the most suitable substrate for B. amyloliquefaciens TKU050 pectinase. Banana peel is a well-known agricultural waste that contains a range of nutrients and bioactivity, including antioxidants [18,19], antimicrobial [20], anti-cancer [21], and enhance α-glucosidase inhibitory in yogurt [22]. Banana peel makes up about 40% of the fruit weight, and the recovery of banana peel for its bioactivity compounds is an essential step toward agricultural sustainability. ...
The utilization of pectin-containing by-products may be useful in a variety of fields. This study aims to establish the processing of pectin-containing by-products to produce pectinases using Bacillus amyloliquefaciens TKU050 strain. In this study, several kinds of agricultural pectin-containing by-products from banana (banana peel), rice (rice bran), orange (orange peel), coffee (spent coffee grounds), and wheat (wheat bran) were utilized to provide carbon sources for the production of a pectinase by B. amyloliquefaciens TKU050. B. amyloliquefaciens TKU050 expressed the highest pectinase productivity (0.76 U/mL) on 0.5% wheat bran-containing medium at 37 °C for four days. A 58 kDa pectinase was purified from the four-day cultured medium fermented under optimized culture conditions with 7.24% of a recovery ratio and 0.51 U/mg of specific activity, respectively. The optimum temperature, optimum pH, thermal stability, and pH stability of the TKU050 pectinase were 50 °C, pH 6, <50 °C, and pH 6–9, respectively. The TKU050 pectinase was inhibited by sodium dodecyl sulfate and Cu2+. The reducing sugar obtained by hydrolyzing banana peel with TKU050 pectinase showed the growth-enhancing effect on the growth of four tested lactic acid bacteria.
Keywords: pectin; pectinase; wheat bran; banana peel; Bacillus amyloliquefaciens; prebiotics
... This type of agricultural waste is usually disposed in landfills and/ or used for agricultural fertilization. Although high-value co-products can be generated from banana cultivation, only a small part of the total volume of banana peels has been explored, mainly as an ingredient rich in dietary fiber [5]. ...
The deconstruction of banana peel for carbohydrate recovery was performed by sequential treatment (acid, alkaline, and enzymatic). The pretreatment with citric acid promoted the extraction of pectin, resulting in a yield of 8%. In addition, xylose and XOS, 348.5 and 17.3 mg/g xylan, respectively, were also quantified in acidic liquor as a result of partial depolymerization of hemicellulose. The spent solid was pretreated with alkaline solution (NaOH or KOH) for delignification and release of residual carbohydrates from the hemicellulose. The yields of xylose and arabinose (225.2 and 174.0 mg/g hemicellulose) were approximately 40% higher in the pretreatment with KOH, while pretreatment with NaOH promoted higher delignification (67%), XOS yield (32.6 mg/g xylan), and preservation of cellulosic fraction. Finally, the spent alkaline solid, rich in cellulose (76%), was treated enzymatically to release glucose, reaching the final concentration of 28.2 g/L. The mass balance showed that through sequential treatment, 9.9 g of xylose, 0.5 g of XOS, and 8.2 g of glucose were obtained from 100 g of raw banana peels, representing 65.8% and 46.5% conversion of hemicellulose and cellulose, respectively. The study of the fractionation of carbohydrates in banana peel proved to be a useful tool for valorization, mainly of the hemicellulose fraction for the production of XOS and xylose with high value applications in the food industry.
... Antioxidant capacity of peel extracts was linked with banana ripening stages wherein the activity increased in ripe fruits while it decreased in overripe fruits. Gurumallesh et al. [87] isolated a novel metalloprotease from banana peel which had high potential to be used as a therapeutic for anti-cancer activity (its mechanism involves breaking down of collagen peptide bonds). ...
Sustainable utilization of agri-food wastes and by-products for producing value-added products (for cosmetic, pharmaceutical or food industrial applications) provides an opportunity for earning additional income for the dependent industrial sector. Besides, effective valorisation of wastes/by-products can efficiently help in reducing environmental stress by decreasing unwarranted pollution. The major focus of this review is to provide comprehensive information on valorisation of agri-food wastes and by-products with focus laid on bioactive compounds and bioactivity. The review covers the bioactives identified from wastes and by-products of plants (fruits, exotic fruits, vegetables and seeds), animals (dairy and meat) and marine (fish, shellfish seaweeds) resources. Further, insights on the present status and future challenges of sustainably utilizing agri-food wastes/by-products for value addition will be highlighted.
... Until now, many researchers have used the waste from banana plants as an alternative to innovative food, alternative energy sources and important materials for water and wastewater treatment. Some researchers use banana peels as antidepressant candy, milk mixture, and animal feed [14,15]. Zhang et al. [16] stated that banana peels can be used as an alternative for battery fillers. ...
The use of organic waste as bio-sorbent has been carried out by many researchers in the world. Furthermore, the utilization of plants for treating wastewater is also commonly found in various environmental applications. Nevertheless, a review of the ability of banana plants ( Musa spp. ) as bio-sorbent to eliminate gaseous pollutants is rarely found and has not been fully understood. In this paper, lignocellulosic biomass from banana plants (bark, stem, leaves, peels, etc) was identified and reviewed. Sorption potential was discussed and taken from various literature which then evaluated to discuss the potential and ability of banana plants as sorbent material for treating gaseous pollutants. Assessment and measurement methods were also discussed to obtain the best sorbent in removing gaseous pollutants. This research was conducted by reviewing scientific articles that discussed the use of lignocellulosic materials derived from banana plants for absorbing various gases. Almost all articles described the manufacture of adsorbents from banana plants that were used to treat wastewater but not many researchers have tried to develop a commercially adsorbent for gas-shaped pollutants. This research is expected to provide essential information for the readers to develop advanced materials used to control environmental pollution especially gaseous pollutants.
... Metalloproteases are one of the most important hydrolytic enzymes used in the industry. There is a vast diversity of metalloproteases in nature, including plant, animal, fungal, agro-industrial and microbial sources, with microbial metalloproteases being the most significant [1,2]. Metalloproteases play a role in the degradation of proteins and are involved in the modulation of cell growth, inflammation, immunity, and hormone processing [3]. ...
Serratiopeptidase, a metalloprotease produced by Serratia marcescens, is produced through a fermentation process using carbohydrates and proteins as carbon and nitrogen sources. However, some byproducts of the silk industry could be an alternative source for serratiopeptidase production. Therefore, the present work is focused on the purification and characterization of a serratiopeptidase produced from the C8 isolate of Serratia marcescens and obtained from a Colombian silkworm hybrid using casein or silkworm pupae. The protease was purified using ultrafiltration, anion-exchange, and size-exclusion chromatography. The purified enzyme showed a molecular weight of ~50 kDa with a purity above 96%, an isoelectric point of ~4.6, optimum pH and temperature of 6 and 50 °C, and stability at 4 °C for one month. The kinetic constants using azocasein as substrate were 0.63 mM (Km), 2,016 μM/min (Vmax), 41.41 s-1 (Kcat), and 6.56 × 107 M-1 s-1 (Kcat/Km). Inhibition by 5 mM EDTA or 1,10-phenanthroline was recovered by adding Zn2+ at the same concentration. Mass spectrometry analysis indicated 94% homology with the sequence of serratiopeptidase produced by the E-15 strain. We purified and characterized a serratiopeptidase produced by the C8 isolate of S. marcescens in a culture medium based on a renewable source from the silk industry.
Activated carbon was produced from banana peel by chemical activation with K2CO3. The impregnated K2CO3 influenced the carbonization behavior at temperatures below 500 °C and caused the specific surface area to increase between 400 and 500 °C. At temperatures above 700 °C, the char was gasified by the K2CO3, so that the specific surface area increased rapidly between 700 and 800 °C and decreased between 800 and 900 °C because of the excess gasification. The activated carbon produced at an impregnation ratio of 1.0 and a carbonization temperature of 800 °C had the maximum specific surface of 2445 m²/g. The amount of adsorbed methane on this activated carbon at 3.0 MPa was 265 mL (s.t.p.)/g, which is almost the same as that of a commercial activated carbon with an almost identical pore structure. The production of such an activated carbon from banana peel using K2CO3 is important because K2CO3 is much easier to handle than KOH.
Novel ‘Banana Grits’ (BG) was prepared from the pulp of unripe banana (Musa AAB) cv. Nendran, and their detailed phytochemical investigation is described. The present work serves as a first report of isolation and identification of glycolipids by exhaustive NMR and HR-ESI-MS analysis from Nendran that include glycolipids viz. monogalactosyldiacylglycerols, digalactosyldiacylglycerols, acyl steryl glycosides, glucocerebrosides, and sterylglycoside. Proximate analysis, estimation of total carotenoid content, and the resistant starch content in BG are also reported. In vitro starch digestion pattern indicated that out of 80% of total starch in BG, 21% is slowly digestible starch (SDS) and 42% is resistant starch (RS). The presence of SDS and RS indicates that they may contribute to gut health and glycaemic control.
In the agroindustry, sieving is a unit operation of great value, this work aims to make a literature review on sieving in cereals, a search equation was carried out in the Scopus database with the keywords sieve, screen, food process, and cereal that resulted in 132 articles and 174 patents. Of the articles, 44 were directly related to sieving and 14 more had something to do with sieving; of the patents, in the last 10 years only 7 were directly related to sieving. To find new trends, raw materials, patent analysis, and information analysis, tables were built with name, year, author, keywords, countries, quartile, journal, relationship with the agroindustry, and purpose. Among the most important conclusions was the application of sieving in raw materials such as Rice, Corn, Wheat, Cotton, Millet, Quinoa, Almonds, Barley, Potato, Yucca, Microorganisms, Oats, Cotton, Protein, Peppers, and Chia Seed. Furthermore, the use of rotating and vibrating sieves was identified, and also their positive effects on the physicochemical, standardization, and classification of raw materials were identified. The different types of equipment or methods focused on sieving, that has been granted use or design patent, were also recognized.
The objective of this study was to evaluate the effect of natural bleaching products on the color, whiteness, and superficial properties of dental enamel. Seventy fragments of bovine teeth were obtained (6mm x 6mm x 2mm). Initial surface roughness (Surfcorder SE1700, Kosakalab), microhardness (HMV-2, Shimadzu), color (EasyShade, VITA), and surface gloss (Micro-Gloss 45º BYK, Gardner) readings were done. Samples were separated into five groups (n=14) according to the treatments used: CT-conventional toothpaste (negative control); CH-charcoal; TU-turmeric; BP-banana peel, and CP16%-16% carbamide peroxide gel (positive control, 4 h/day for 14 days), and then brushed for 560 cycles (T1) and 1200 cycles (T2), equivalent to 14 and 30 days of brushing. New measurements were performed after T1 and T2. The whiteness index for dentistry change (∆WID) and Weight loss (Wl) were calculated. CP16% demonstrated the highest (p<.05) color change (ΔE00) and ∆WID (2-way ANOVA, Bonferroni, p<.05). Surface gloss alterations were lower for TU, CP16%, and BP. CT and CH increased surface roughness (p<.05). CP16% decreased enamel microhardness. CH presented medium abrasiveness, and CT and TU, low abrasiveness. The popular bleaching products were not efficient for tooth whitening. Furthermore, brushing with charcoal increased the enamel surface roughness, and CP16% decreased enamel microhardness over time.
Due to the increasing world population and ever developing technology, the need and demand for energy are increasing day by day. In parallel with this, the tendency to use renewable alternative energy sources instead of limited fossil fuel reserves is increasing worldwide. Lignocellulosic biomasses which are abundant in nature with renewable energy potential are preferred in biofuel production. These raw biomaterials are transformed into forms that can be used in biofuel production processes by various pretreatment techniques. The physical and chemical methods commonly used in the pretreatment of the substrate have some limitations. However, microbial methods for hydrolysis of biomass are quite remarkable. In this study, we focused on the pretreatment of biomass and microbial enzymes used in biofuel production process.
The fruit processing industry generates large amounts of wastes (pomace, seeds, peels) that causes negative environmental impact with considerable treatment expenses. Nevertheless, various studies demonstrated that these by-products are still rich in bioactive compounds, especially dietary fibres and phenolic compounds, thus leading to significant chemical, physical and biological properties. These characteristics make fruits by-products a good source for new supplements in food products having important effect on intestinal function. Thus, the aim of this review is to evaluate the different bioactive compounds isolated from fruit by-products and to analyse their application in various formulations for the food and nutraceutical industries. In consideration of the biological properties of these compounds, their role in the functioning and action on intestine and gut flora was discussed.
This study was focused on investigating the anticancer potential of indole-3-carbinol (I3C) against osteosarcoma MG-63 and U2OS cells. A wound healing assay indicated that IC3 inhibited migration of MG-63 and U2OS cells. MTT, WST-1, and colony formation assays revealed that treatment of MG-63 and U2OS cells with I3C decreased cell viability. Fluorescence-activated cell sorting (FACS) analysis showed that I3C induced apoptosis in a dose- and time-dependent manner in MG-63 and U2OS cells. Moreover, via terminal deoxynucleotidyl transferase- (TdT-) mediated dUTP-biotin nick-end labeling (TUNEL) assay, we detected that I3C induced DNA fragmentation. Western blotting demonstrated that activated forms of caspase-3, caspase-7, and caspase-9, as well as poly (ADP-ribose) polymerase (PARP) were increased in MG-63 and U2OS cells, following treatment with I3C. Furthermore, protein expression levels of FOXO3, Bax, and Bim extra-large form were increased while those of Akt, JNK, p38, phosphorylated ERK, and Bcl-xL were decreased by I3C treatment in MG-63 and U2OS cells. Thus, the study indicates that I3C may induce apoptosis in human osteosarcoma MG-63 and U2OS cells via the activation of apoptotic signaling pathways by FOXO3.
Protease occurs naturally in all organisms and is an essential constituent for all the existing live forms. Microorganisms such as bacteria and fungi and yeast are the main source of protease enzyme. They act as an important industrial enzyme occupying for about 60% of total enzyme market. In this study, protease was isolated from various leaves such as Coriandrum sativum, Nicotiana tobaccum, Murraya koenigii, and Moringa oleifera was partially purified. Then their specific activity and optimum pH were checked. Among the four plant species, the protease activity was found to be more in Nicotiana tobaccum (5.6 units/mg of protein) followed by Moringa oleifera (2.46 units/mg of protein), Murraya koenigii (2.02 units/mg of protein) and least in Coriandrum sativum (1.56 units/mg of protein). The optimum pH was found to be 7.2 for all the samples.
Protease plays an important role in the management of defense mechanism of living organisms. This review presents a comprehensive and exhaustive account of plant, animal and microbial enzymes with special reference to proteolytic properties. A large number of biological sources of proteases described in this review
clearly demonstrate the importance of plants and microbial proteases in applied and industrial uses. Proteases have potential applications in food, dairy, detergent, leather, alcoholic beverages, brewing, meat, pharmaceutical and photographic industries. The research work on protease has been going on since seven decades but no exhaustive review on protease from last eleven years is available in the literature. This review attempts to focus on some of the difficulties observed in the earlier work and tried to find out possible solutions and bridging up this gap by introducing recent information regarding proteinases. This review provides information on diverse groups of proteases with respect to scientific name, family, purification scheme, protease inhibitors, physiological functions, also industrial applications, sequence homology and the future scope of protease enzymes.
BackgroundUrease, one of the highly efficient known enzymes, catalyzes the hydrolysis of urea into ammonia and carbon dioxide. The present study aimed to extract urease from pea seeds (Pisum Sativum L). The enzyme was then purified in three consequence steps: acetone precipitation, DEAE-cellulose ion-exchange chromatography, and gel filtration chromatography (Sephacryl S-200 column).ResultsThe purification fold was 12.85 with a yield of 40%. The molecular weight of the isolated urease was estimated by chromatography to be 269,000 Daltons. Maximum urease activity (190 U/g) was achieved at the optimum conditions of 40°C and pH of 7.5 after 5 min of incubation. The kinetic parameters, K
m
and V
max
, were estimated by Lineweaver-Burk fits and found to be 500 mM and 333.3 U/g, respectively. The thermodynamic constants of activation, ΔH, E
a
, and ΔS, were determined using Arrhenius plot and found to be 21.20 kJ/mol, 23.7 kJ/mol, and 1.18 kJ/mol/K, respectively.ConclusionsUrease was purified from germinating Pisum Sativum L. seeds. The purification fold, yield, and molecular weight were determined. The effects of pH, concentration of enzyme, temperature, concentration of substrate, and storage period on urease activity were examined. This may provide an insight on the various aspects of the property of the enzyme. The significance of extracting urease from different sources could play a good role in understanding the metabolism of urea in plants.
A bacterial strain producing both a protease and an esterase was isolated from the seed of Tamarindus indica. The isolate was identified as Lysinibacillus fusiformis AU01 by phylogenetic analysis of the 16S rDNA sequence. The isolate produced an extracellular protease and an intracellular esterase simultaneously under the same culture conditions. Statistical methods, i.e., Plackett-Burman followed by response surface methodology (RSM), were applied to optimize media components and culture conditions in 50 mL shake flask cultures. Culturing in shake flasks with the optimized medium resulted in a 6-fold and a 3.5-fold increase in protease and esterase production, respectively, when compared to nutrient medium. The productivity of the protease and esterase was increased to 75 U/mL and 370 U/mL when cultivated under controlled conditions in a 3-L bioreactor. The extracellular protease was purified to 34.6 fold with 38.8 % recovery and the molecular mass of the enzyme was found to be 48 kDa. The partial amino acid sequence of the protease was determined by MALDI-TOF-MS analysis. The optimum temperature and pH for protease activity were found to be pH 9.0 and 40 °C. The inhibition of purified protease by EDTA and PMSF confirmed that the enzyme belonged to the family of serine metalloproteases. The enzyme was found to be stable in the presence of some hydrophobic and hydrophilic solvents. The ability of AU01 protease to dissociate monolayer cells for sub-culturing adherent cell lines was also investigated.
Industrial and household catalysis becomes more and more dependent on enzymes. This is not surprising since enzymes are able to catalyze all kinds of chemical reactions. Enzymes with the desired activity under industrial conditions can be obtained by optimizing process conditions and by protein engineering. The use of enzymes frequently results in many benefits that cannot be obtained with traditional chemical treatment. These often include higher product quality and lower manufacturing cost, less waste and reduced energy consumption. Key factors driving the market growth include new enzyme technologies endeavoring to enhance cost efficiencies and productivity, and growing interest among consumers in substituting petrochemical products with other organic compounds such as enzymes. Other factor propelling market growth includes surging demand from textile manufacturers, animal feed producers, detergent manufacturers, pharmaceutical companies, bioethanol producers and cosmetics vendors. The present paper aims to provide a review on industrial enzymes, highlighting on recent scientific advances, current applications in diverse industrial sectors and global market.
Proteolytic enzymes (also termed peptidases, proteases and proteinases) are capable of hydrolyzing peptide bonds in proteins. They can be found in all living organisms, from viruses to animals and humans. Proteolytic enzymes have great medical and pharmaceutical importance due to their key role in biological processes and in the life-cycle of many pathogens. Proteases are extensively applied enzymes in several sectors of industry and biotechnology, furthermore, numerous research applications require their use, including production of Klenow fragments, peptide synthesis, digestion of unwanted proteins during nucleic acid purification, cell culturing and tissue dissociation, preparation of recombinant antibody fragments for research, diagnostics and therapy, exploration of the structure-function relationships by structural studies, removal of affinity tags from fusion proteins in recombinant protein techniques, peptide sequencing and proteolytic digestion of proteins in proteomics. The aim of this paper is to review the molecular biological aspects of proteolytic enzymes and summarize their applications in the life sciences.
A neutral protease was detected in the culture medium of Streptomyces microflavus isolated from some Egyptian soils. The enzyme was purified by precipitation with ammonium sulphate and gel filtration on Sephadex G-75. The optimal pH and temperature for catalytic activity of protease was pH 7 and 40 0C respectively. Calcium and manganese stimulated protease activity while Ag+ inhibited the enzyme. The proteolytic activity of protease was strongly inhibited by 0.8 mM of Para chloromercuribenzoic acid (P. CMBA) and phenyl methyl sulfonyl floride (PMSF).
Five different mutant strains were developed from the wild strain of Saccharomyces cerevisiae (MTCC No.287) using UV irradiation technique by varying the exposure timings. All the mutant cultures were used for ethanol production using banana peel as a substrate in a batch fermenter. The effect of temperature, pH and initial substrate concentration on ethanol production were studied and optimized. The mutant strain 4 gave a maximum ethanol production of 9 g/L under identical conditions. The conditions were optimized for mutant strain 4 and a temperature of 33 °C, pH 4.5 and initial substrate concentration 10%(w/v) were found to be optimum. The kinetics of ethanol production using mutant strain 4 under optimum conditions was studied and modeling was attempted using different models. Monod model for growth kinetics and Leudeking-Piret model for product formation kinetics were found to represent the experimental data very well.
A protease enzyme was isolated and partially purified from the pulp of Thaumatococcus daniellii fruit
by gel filtration on sephadex G-75 followed by ion-exchange column chromatography on DEAEcellulose.
The enzyme showed a specific activity of 4.75 × 10-1 unit/mg protein and 6.93 × 10-1 unit/mg
protein, respectively after each purification procedure. The purified enzyme had a Km and Vmax of 2.0 ×
10-4 M and 1.53 mol/min, respectively, using casein as substrate. The enzyme had an optimum
temperature of 35°C and functioned best at pH 7.0 with some residual activity at alkaline pH.
This review presents a brief overview of the general categories of commercially used proteases, and critically surveys the successful strategies currently being used to improve the properties of proteases for various commercial purposes. We describe the broad application of proteases in laundry detergents, food processing, and the leather industry. The review also introduces the expanding development of proteases as a class of therapeutic agents, as well as highlighting recent progress in the field of protease engineering. The potential commercial applications of proteases are rapidly growing as recent technological advances are producing proteases with novel properties and substrate specificities.
Musa sapientum peels were analysed for minerals, nutritional and anti - nutritional contents. The result of mineral content indicate the concentrations (mg/g) of potassium, calcium, sodium, iron, manganese, bromine, rubidium, strontium, zirconium and niobium to be 78.10, 19.20, 24.30, 0.61, 76.20, 0.04, 0.21, 0.03, 0.02 and 0.02 respectively. The percentage concentrations of protein, crude lipid, carbohydrate and crude fibre were 0.90, 1.70, 59.00 and 31.70 respectively. The results indicate that if the peels are properly exploited and process, they could be a high-quality and cheap source of carbohydrates and minerals for livestock.
Banana serves as an ideal and low cost food source for developing countries where most of the population rely mostly on bananas for food. Banana plant parts are useful as insecticide, antioxidant, colour absorber, in preparation of various functional foods, wine, alcohol, biogas, cattle feed etc. This review discusses usefulness of banana fruits, peel, leaves, pseudostem, sheath, pith and male bud, and prospects of using these materials in industry.
Proteases break peptide bonds. In the lab, it is often necessary to measure and/or compare the activity of proteases. Sigma's non-specific protease activity assay may be used as a standardized procedure to determine the activity of proteases, which is what we do during our quality control procedures. In this assay, casein acts as a substrate. When the protease we are testing digests casein, the amino acid tyrosine is liberated along with other amino acids and peptide fragments. Folin and Ciocalteus Phenol, or Folin's reagent primarily reacts with free tyrosine to produce a blue colored chromophore, which is quantifiable and measured as an absorbance value on the spectrophotometer. The more tyrosine that is released from casein, the more the chromophores are generated and the stronger the activity of the protease. Absorbance values generated by the activity of the protease are compared to a standard curve, which is generated by reacting known quantities of tyrosine with the F-C reagent to correlate changes in absorbance with the amount of tyrosine in micromoles. From the standard curve the activity of protease samples can be determined in terms of Units, which is the amount in micromoles of tyrosine equivalents released from casein per minute.
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Urease purified from pigeonpea seeds was immobilized on gelatin beads via cross-linking with glutaraldehyde. The maximum immobilization (75%) was observed at 30 mg/ml gelatin, 0.414 mg of enzyme/bead, 1% (v/v) glutaraldehyde and 4 degrees C. Beads stored in 50 mM Tris/acetate buffer (pH 7.3) at 4 degrees C showed a half-life of 240 days and there was practically no leaching of enzyme (less than 2%) over a period of 30 days. These beads can be reused more than 30 times (with 24 h intervals) without much loss of enzyme activity (i.e. less than 11%). The immobilized urease showed a shift in its optimum pH from 7.3 to 6.5 in Tris/acetate buffer. Optimum temperature also shifted from 47 to 65 degrees C compared with the soluble enzyme. Gelatin-immobilized pigeonpea urease had a higher K(m) (8.3 mM) than that of the soluble enzyme (3.0 mM). The time-dependent temperature inactivation pattern was also found to change from biphasic to monophasic kinetics. The immobilized beads were used for the preparation of a new urea biosensor with a response time of less than 2 min. At least 14 samples of urea can be measured with this biosensor within an hour. The beads, as well as the biosensor, were used to analyse the urea content in clinical samples from the local clinical pathology laboratories. The results obtained with the biosensor were strikingly similar to those obtained with the various commonly employed biochemical/autoanalyzer(R) methods used. These immobilization studies also have a potential role in haemodialysis machines that maintain the urea level in kidney patients and in the construction of a portable/wearable kidney. The easy availability of the pigeonpea urease, the ease of its immobilization on gelatin and a significantly lower cost of the urease described in the present study makes it a suitable product for future applications in therapeutics and diagnostics.
This work examines the influence of various process parameters (like sodium alginate concentration, calcium chloride concentration, and hardening time) on papain entrapped in ionotropically cross-linked alginate beads for stability improvement and site-specific delivery to the small intestine using neural network modeling. A 3(3) full-factorial design and feed-forward neural network with multilayer perceptron was used to investigate the effect of process variables on percentage of entrapment, time required for 50% and 90% of the enzyme release, particle size, and angle of repose. Topographical characterization was conducted by scanning electron microscopy, and entrapment was confirmed by Fourier transform infrared spectroscopy and differential scanning calorimetry. Times required for 50% (T(50)) and 90% (T(90)) of enzyme release were increased in all 3 of the process variables. Percentage entrapment and particle size were found to be directly proportional to sodium alginate concentration and inversely proportional to calcium chloride concentration and hardening time, whereas angle of repose and degree of cross-linking showed exactly opposite proportionality. Beads with >90% entrapment and T(50) of <10 minutes could be obtained at the low levels of all 3 of the process variables. The inability of beads to dissolve in acidic environment, with complete dissolution in buffer of pH >or=6.8, showed the suitability of beads to release papain into the small intestine. The shelf-life of the capsules prepared using the papain-loaded alginate beads was found to be 3.60 years compared with 1.01 years of the marketed formulation. It can be inferred from the above results that the proposed methodology can be used to prepare papain-loaded alginate beads for stability improvement and site-specific delivery.
Proteases are a group of large complex enzyme molecules that perform highly focused proteolysis functions. A vast quantity of the protease enzymes is predominantly sourced from microbial fermentation process, although proteases tend to natively present in plant, animals and humans. Proteases possess a pervasive importance in medical and pharmaceutical sector, because of its enriched specificity towards biomolecules. They are also actively encompassed in regulating certain physiological pathways. A distinct territory of human disorders is treated by substrate specific proteases. Enormous numbers of catalytic activities in habitual metabolism process of a living organism are protease dependent. Pilot scale researches and product development in industrial biotechnology sectors are wholly based on any one of the protease enzymes. The applications of the protease enzymes and its economic benefits of being an eco-friendly material are far-reaching. This review presents a brief overview on the classification and sources of various types of proteases. We describe the essential evidences of role of protease in different sectors. The proteases could be a potential relieves to harmful synthetic chemicals in distinctive industrial processes and thus gains global perception.
The crude fraction extracted at pH 6.0 from sprouting potato tubers (pH 6.0 fraction) hydrolyzed casein and BANA at pH 6.0. This pH 6.0 fraction contained not only caseinase activity but also gelatinase activity, detected by active staining of PAGE-gel with gelatin, as endopeptidases, and both activities increased during sprouting of tubers. This endopeptidase, also active on Azocolase, had an optimum pH at pH 6.0, whereas the crude fraction extracted at pH 6.0 from fresh potato tubers contained little endopeptidase activities in the whole pH range. Inhibition by monoiodoacetate or antipain indicated this endopeptidase to be a cysteine protease.
An alkaline proteinase (STAP) was produced from strain TN650 isolated from a Tunisian off-shore oil field and assigned as Streptomyces koyangensis strain TN650 based on physiological and biochemical properties and 16S rRNA gene sequencing. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis revealed that the purified enzyme was a monomer with a molecular mass of 45125.17-Da. The enzyme had an NH2-terminal sequence of TQSNPPSWGLDRIDQTTAFTKACSIKY, thus sharing high homology with those of Streptomyces proteases. The results showed that this protease was completely inhibited by phenylmethanesulfonyl fluoride (PMSF), diiodopropyl fluorophosphates (DFP), and partially inhibited by 5,5-dithio-bis-(2-nitro benzoic acid) (DTNB), which strongly suggested its belonging to the serine thiol protease family. Using casein as a substrate, the optimum pH and temperature values for protease activity were pH 10 and 70°C, respectively. The protease was stable at pH 7-10 and 30-60°C for 24h. STAP exhibited high catalytic efficiency, significant detergent stability, and elevated organic solvent resistance compared to the SG-XIV proteases from S. griseus and KERAB from Streptomyces sp. AB1. The stap gene encoding STAP was isolated, and its DNA sequence was determined. These properties make STAP a potential candidate for future application in detergent formulations and non-aqueous peptide biocatalysis.
Copyright © 2015. Published by Elsevier B.V.
Plant-derived compounds have played an important role in the development of several clinically useful anti-cancer agents. These include vinblastine, vincristine, the camptothecin derivatives, topotecan and irinotecan, etoposide, and paclitaxel (Taxol®). Several promising new agents are in clinical development based on selective activity against cancer-related molecular targets, including flavopiridol and Combretastatin A4 phosphate. Recently, plants have yielded several agents showing anti-AIDS activity, and one of these, (+)-calanolide A, is in clinical development.
The presence of a protease in Artocarpus integer leaves, which are traditionally used as a meat tenderiser, was verified by the presence of a band at 69kDa, using caseinolytic zymography. Purification by temperature phase partitioning with Triton X-114, ammonium sulphate precipitation and gel filtration chromatography yielded a preparation with a 12-fold increase in enzyme purity and a final specific activity of 76.67U/mg. The cysteinic nature of this enzyme was confirmed through inhibition of enzyme activity by E-64 and iodoacetamide and enhancement of activity by cysteine and 2-mercaptoethanol. The protease retained 70% of its activity over a broad pH range (pH 6–12), with optimal activity recorded at pH 10 and 40°C. The enzyme was stable at temperatures up to 70°C, with 80% of its activity intact. Addition of 5mM Ca2+ stimulated enzyme activity and a kinetic study of the enzyme yielded Km and Vmax values of 0.304mg/mL and 0.735mg/mL/min, respectively.
This article reviews background on proteases and their functions, their physiological significance in skin, and the potential implications of incorporating specific proteases and protease blends into dermatological products, including skin care formulations. The history of protease blend formulations used in wound model studies and for other disorders is reviewed. In vitro data with use of a specific 3-protease blend with evaluation of the impact on various skin proteins and peptides is also discussed in this article.
Lipase production (8.02±0.24U/ml) by the yeast Aureobasidium pullulans HN2.3 isolated from sea saltern was carried by using time-dependent induction strategy. The lipase in the supernatant of the yeast cell culture was purified to homogeneity with a 3.4-fold increase in specific lipase activity as compared to that in the supernatant by ammonium sulfate fractionation, gel filtration chromatography and anion-exchange chromatography. According to the data on SDS polyacrylamide gel electrophoresis, the molecular mass of the purified enzyme was estimated to be 63.5kDa. The optimal pH and temperature of the purified enzyme were 8.5 and 35°C, respectively. The enzyme was greatly inhibited by Hg2+, Fe2+ and Zn2+. The enzyme was strongly inhibited by phenylmethanesulphonyl fluoride, not inhibited by ethylene diamine tetraacetic acid (EDTA), but weakly inhibited by iodoacetic acid. It was found that the purified lipase had the highest hydrolytic activity towards peanut oil.
Banana must was treated with pectinase and α ‐amylase to hydrolyze pectin and starch prior to its use to produce a wine product. The synergistic activities of the enzymes enhanced hydrolysis of the complex carbohydrates. A decrease of 55% in the viscosity and a 2.7‐fold increase in the amount of extracted juice were obtained after incubating with 0.05% (w/w) of pectinase at 40C for 2 h, followed by treating with 0.05% (w/w) of α ‐amylase at 50C for 3 h. A 15 and 39% increase in total soluble sugars and reducing sugars in extracted juice were achieved, respectively. Enzyme‐treated banana must was diluted with four volumes of water and then fermented by yeast to produce banana wine. The pretreatment of banana with enzymes before wine fermentation resulted in a higher level of reducing sugars than that of the control (nonenzyme‐treated banana wine) during fermentation. The clarity of the enzyme‐treated banana wine was also fourfold higher than that of the control at 25 days of fermentation. The concentrations of total soluble solids, total soluble sugars, and alcohol in the enzyme‐treated banana wine and the control have no significant differences.
PRACTICAL APPLICATIONS
Banana fruits contain high nutrition sources of carbohydrate, minerals and vitamins. Banana‐based wine has a good flavor and can be considered ahealthy alcoholic beverage. However, polymeric carbohydrates like pectin and starch in banana cause the turbidity and viscosity of the wine and make the clarification process harder. Application of pectinase and α ‐amylase that affect the quality of wine is important for improving the process of banana wine production. This study highlights the effectiveness of pectinase and α ‐amylase treatments on banana wine production such as increasing the banana juice extraction yield, decreasing the viscosity of banana must, forming simple sugars and oligosaccharides to facilitate yeast growth during fermentation and improving the clarification process.
1. Vicilin peptidohydrolase, the major endopeptidase present in the cotyledons of bean seedlings, has been purified to homogeneity, as judged by polyacrylamide gel electrophoresis and isoelectrofocusing, from the cotyledons of mung bean [Vigna radiata (L.) Wilczek] seedlings.
2. The protease has an apparent molecular weight of 23 000 as estimated by gel filtration and polyacrylamide gel electrophoresis, an isoelectric point of 3.75 and belongs to the class of the sulfhydryl proteases as judged by the effect of various protease inhibitors on the activity of the enzyme.
3. The purified enzyme readily hydrolyzes synthetic esters between amino acids and p-nitrophenol coupled to N-carbobenzoxy and N-tert-butoxycarbonyl residues. Esters with asparagine and glutamine residues are hydrolyzed most readily.
4. The purified enzyme readily degrades vicilin, the principal reserve protein present in mung bean seeds. Degradation is optimal at pH 5.1. The various polypeptides of vicilin are gradually digested to smaller polypeptides and oligopeptides. Vicilin peptidohydrolase and carboxypeptidase cooperate in the complete digestion of vicilin with the resulting release of amino acids.
ZYMV-AGII (zucchini yellow mosaic virus-AGII) is a recombinant nonpathogenic potyvirus-based vector system for the expression of foreign genes in cucurbit plants and their edible fruits, including squash, cucumber, melon, watermelon, and pumpkin. MAP30 (Momordica anti-HIV protein, 30 kDa) and GAP31 (Gelonium anti-HIV protein 31 kDa) are multifunctional plant proteins with activity against HIV-1 virus. These proteins are also effective against other viruses, tumor cells, and microbes. We report here the production and characterization of biologically active MAP30 and GAP31 in squash plant by expression of their genes using the ZYMV-AGII vector. Recombinant expressed MAP30 and GAP31 exhibit comparable antiviral, antitumor, and antimicrobial activities as their counterparts from their original plant sources, with EC(50)s in the ranges of 0.2-0.3 nM for HIV-1. These results demonstrate for the first time the amplification and production of therapeutic proteins, MAP30 and GAP31, in common vegetables. This provides valuable alternative food sources of these antiviral, antitumor, and antimicrobial agents for therapeutic applications.
Urease has been purified from the dehusked seeds of pigeonpea (Cajanus cajan L.) to apparent electrophoretic homogeneity with approximately 200 fold purification, with a specific activity of 6.24 x10(3) U mg(-1) protein. The enzyme was purified by the sequence of steps, namely, first acetone fractionation, acid step, a second acetone fractionation followed by gel filtration and anion-exchange chromatographies. Single band was observed in both native- and SDS-PAGE. The molecular mass estimated for the native enzyme was 540 kDa whereas subunit values of 90 kDa were determined. Hence, urease is a hexamer of identical subunits. Nickel was observed in the purified enzyme from atomic absorption spectroscopy with approximately 2 nickel ions per enzyme subunit. Both jack bean and soybean ureases are serologically related to pigeonpea urease. The amino acid composition of pigeonpea urease shows high acidic amino acid content. The N-terminal sequence of pigeonpea urease, determined up to the 20th residue, was homologous to that of jack bean and soybean seed ureases. The optimum pH was 7.3 in the pH range 5.0-8.5. Pigeonpea urease shows K(m) for urea of 3.0+/-0.2 mM in 0.05 M Tris-acetate buffer, pH 7.3, at 37 degrees C. The turnover number, k(cat), was observed to be 6.2 x 10(4) s(-1) and k(cat)/K(m) was 2.1 x 10(7) M(-1) s(-1). Pigeonpea urease shows high specificity for its primary substrate urea.
The pepstatin A sensitive acidic proteolytic activity of total protein extracts of buckwheat seeds has been analyzed in developing, mature, and germinating seeds by activity measurements as well as by electrophoretic and immunochemical techniques. Immunoblot analysis using cross-reactive antibodies raised against barley phytepsin suggested that specific proteolytic activity could be attributed to a 47 kDa heterodimeric polypeptide, composed of two subunits: 31 and 16 kDa polypeptides. The analysis of time course expression revealed that the 47 kDa heterodimer accumulated during seed maturation starting from 12 days after pollination and was also present at the beginning of germination. Milk-clotting activity of this proteinase was also indicated.
Plant-derived compounds have been an important source of several clinically useful anti-cancer agents. These include vinblastine, vincristine, the camptothecin derivatives, topotecan and irinotecan, etoposide, derived from epipodophyllotoxin, and paclitaxel (taxol A number of promising new agents are in clinical development based on selective activity against cancer-related molecular targets, including flavopiridol and combretastin A4 phosphate, while some agents which failed in earlier clinical studies are stimulating renewed interest.
A protein, with a novel N-terminal amino acid sequence and a molecular mass of 30 kDa, was purified from fresh Smilax glabra rhizomes by adsorption on DEAE-cellulose, CM-cellulose, Con A-Sepharose, and Mono S, and by fast protein liquid chromatography-gel filtration on Superdex 75. The protein, designated as smilaxin, stimulated uptake of [methyl-3H]thymidine by murine splenocytes, peritoneal macrophages, and bone marrow cells, and production of nitric oxide by peritoneal macrophages. It inhibited uptake of [methyl-3H]thymidine by MBL2 and PU5 tumor cells but not uptake by S180 and L1210 cells. Smilaxin augmented glucose uptake into rat adipose tissue. It attenuated the activity of HIV-1-reverse transcriptase with an IC50 of 5.6 microM. However, it did not display hemagglutinating, antifungal or translation-inhibitory activities, indicating that it is not a lectin, an antifungal protein, or a ribosome-inactivating protein.
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