Debittering Of Juices Research Articles

Amination of naringinase to improve citrus juice debittering using a catalyst immobilized on glyoxyl-agarose

A naringinase complex was chemically aminated prior to its immobilization on glyoxyl-agarose to develop a robust biocatalyst for juice debittering. The effects of amination on the optimal pH and temperature, thermal stability, and debittering performance were analyzed. Concentration of amino groups on catalysts surface increased in 36 %. Amination reduced the β-glucosidase activity of naringinase complex; however, did not affect optimal pH and temperature of the enzyme and it favored immobilization, obtaining α-l-rhamnosidase and β-d-glucosidase activities of 1.7 and 4.2 times the values obtained when the unmodified enzymes were immobilized. Amination favored the stability of the immobilized biocatalyst, retaining 100 % of both activities after 190 h at 30 °C and pH 3, while its non-aminated counterpart retained 80 and 52 % of α-rhamnosidase and β-glucosidase activities, respectively. The immobilized catalyst showed a better performance in grapefruit juice debittering, obtaining a naringin conversion of 7 times the value obtained with the non-aminated catalyst.

Enzymatic debittering of citrus juices: Optimization, modeling, and characterization of naringinase production from marine Bacillus subtilis strain BSnari

The bitterness of citrus juices is a major obstacle to their industrial acceptance. Bitterness can be reduced using the naringinase enzyme. This study investigated two approaches for producing and purifying naringinase. Bacillus subtilis strain Bsnari was isolated from the Red Sea with accession number OQ576096 and had the highest naringinase productivity (7.09 ± 0.09 U/ml) using sucrose (1.5%) as carbon source, 0.6% w/v of citrus peel powder and 1% w/v of soy bean meal at pH 7.0 and 37 °C. Box-Behnken design has been successfully used for statistical modeling of naringinase production by Bacillus subtilis. Naringinase was partially purified using ammonium sulfate and gel filtration, resulting in a specific activity of 215.106 U/mg, a purification fold of 8.95, and a molecular weight of 73 KDa. Enzymatic hydrolysis reduced bitterness by around 33–36%, with optimal debittering after 4 h at 40–50 °C. A new method for processing citrus juice debittering was proposed using α-l-rhamnosidase and β-d-glucosidase encoded by the genes rhaA and bglA, respectively. The protein structures of β-l-rhamnosidase and β-d-glucosidase were also investigated. The topology of the 3D structure of α-l-rhamnosidase and β-d-glucosidase proteins of Bacillus sp. Predicted and aligned against other bacterial species. These findings provide insights into efficient naringinase production and purification for use in various applications, with implications for commercial production in food and pharmaceutical industries.

Grapefruit Debittering by Simultaneous Naringin Hydrolysis and Limonin Adsorption Using Naringinase Immobilized in Agarose Supports.

Naringin and limonin are the two main bitter compounds of citrus products such as grapefruit juice. The aim of this investigation was to evaluate the reduction in both bitter components simultaneously using a combined biochemical and physical approach. The proposed strategy was based on the use of heterofunctional supports with glyoxyl groups that allow for the covalent immobilization of naringinase, which hydrolyses naringin and alkyl groups that allow for the adsorption of limonin. The supports were butyl-glyoxyl agarose (BGA) and octyl-glyoxyl agarose (OGA), which were characterized in terms of aldehyde group quantification and FTIR analysis. The optimal pH and temperature of free and immobilized enzymes were assessed. The maximum enzyme loading capacity of supports was analyzed. Debittering of grapefruit juice was evaluated using soluble enzyme, enzyme-free supports, and immobilized catalysts. Enzyme immobilized in BGA reduced naringin and limonin concentrations by 54 and 100%, respectively, while the use of catalyst immobilized in OGA allowed a reduction of 74 and 76%, respectively, obtaining a final concentration of both bitter components under their detection threshold. The use of OGA biocatalyst presented better results than when soluble enzyme or enzyme-free support was utilized. Biocatalyst was successfully applied in juice debittering in five repeated batches.

Immobilization of naringinase on asymmetric organic membranes: Application for debittering of grapefruit juice

An enzymatic membrane reactor (EMR) is assembled for the immobilization of naringinase on a polyethersulfone ultrafiltration membrane, based on fouling-induced method. The effects of molecular weight cut-off, membrane configuration, applied pressure, enzyme concentration and pH are studied in terms of permeate rate, immobilization efficiency, and biocatalytic conversion. The 10 kDa membrane operating in reverse mode, 0.2 MPa, 0.3 gL−1 of enzyme in acetate buffer at pH 5 and cross-linking with 0.25% glutaraldehyde showed the highest naringin conversion (73%). It was determined that the intermediate pore blocking model was the predominant fouling mechanism for the enzymatic immobilization. The EMR was applied for debittering of grapefruit juice, achieving a conversion of naringin below bitterness threshold and maintaining the antioxidant capacity of the juice. Furthermore, the biocatalytic activity of immobilized enzyme was retained at a high level at least during three consecutive reaction runs, with overnight storage at 4 °C after each run. Industrial relevanceThe potential of membrane technologies in the juice industries is widely recognized today. The development of EMR with naringinase activity is an attractive option for reducing bitterness that could replace current techniques, due to its high specificity and effectiveness, possibility of repeated and continuous use, and in order to retain the properties of juice as much as possible. The research therefore represents an advance in the application of biocatalytic membranes as technological alternative for juice debittering.

The effects of α‐L‐rhamnosidase, β‐D‐glucosidase, and their combination on the quality of orange juice

The naringinase, a desirable candidate for citrus juice debittering, consists of α-L-rhamnosidase and β-D-glucosidase. This study aimed to elucidate the effects of α-L-rhamnosidase, β-D-glucosidase, and their combination on naringin, polyphenols, antioxidant activity, and aroma quality. The α-L-rhamnosidase and the two enzymes combination reduced the naringin concentration from 356.33 to 10.52 and 4.17 mg/L, respectively. The α-L-rhamnosidase, β-D-glucosidase, and their combination significantly increased the antioxidant activity. The unpleasant cooking note was significantly weakened and the floral note was increased after three treatments. The combinational treatment increased the sweet note greatly. The substrate competition between α-L-rhamnosidase and β-D-glucosidase may help to explain the inconsistencies of the polyphenols, antioxidant activity, and aroma quality after single enzyme or combined enzymes treatments. The results indicated that taste, antioxidant activity, and aromatic quality were well improved by the combined treatment than either α-L-rhamnosidase or β-D-glucosidase treatment, providing an attractive reference for debittering orange juice. Practical applications The work showed that the combined treatment of α-L-rhamnosidase and β-D-glucosidase was better to remove the bitter taste of citrus juice than the solo treatment with α-L-rhamnosidase or β-D-glucosidase in regarding with taste, antioxidant activity, and aromatic quality. It helps to better the quality of citrus juice with enzymatic treatment.

History of cyclodextrin-based polymers in food and pharmacy: a review.

Cyclodextrins are glucose macrocycles whose inclusional capabilities towards non-polar solutes can be modulated with the help of other macrostructures. The incorporation of cyclodextrin moieties into larger structures produces five types of new materials: crosslinked networks, functionalized chains, amphiphilic cyclodextrins, polyrotaxanes and nanocomposites. This review presents crosslinking and grafting to prepare covalently-attached cyclodextrins, and applications in the food and pharmaceutical sectors, from an historical point of view. In food science, applications include debittering of juices, retention of aromas and release of preservatives from packaging. In biomedical science, cyclodextrin polymers are applied classically to drug release, and more recently to gene delivery and regenerative medicine. The remarkable points are: 1) epichlorohydrin and diisocyanates have been extensively used as crosslinkers since the 1960s, but during the last two decades more complex cyclodextrin polymeric structures have been designed. 2) The evolution of cyclodextrin polymers matches that of macromolecular materials with regard to complexity, functionality and capabilities. 3) The use of cyclodextrin polymers as sorbents in the food sector came first, but smart packaging is now an active challenge. Cyclodextrins have also been recently used to design treatments against the coronavirus disease 2019 (COVID-19).

Isolation and characterization of naringinase enzyme and its application in debittering of Pomelo juice ( Citrus grandis ): A comparative study with macroporous resin

The naringinase enzyme was isolated from Citrus macroptera peel and pomace using Aspergillus niger for immobilization and debittering of juice. A comparative study of the effect of debittering using immobilized enzyme and resin on the physicochemical and phytochemical properties of the juice were carried out. The optimum hydrolytic conditions and Michaelis constant for an immobilized enzyme were observed at pH 4, 70℃, and 16.75 μg/ml, respectively. Adsorption of naringin onto resin followed pseudo‐first‐order with a regression coefficient of 0.92. Naringin content was reduced up to 73.28% in 4 min by resin and 79.76% in 120 min by the enzyme. We reutilized resin and immobilized enzyme effectively to debitter the juice up to seven cycles. Treatment for less than 2 min and 90–120 min using resin and enzyme, respectively, could retain the sizable amount of bioactive components in juice and immobilization retained 62% enzymatic activity during 2 months of storage at 4°C. PRACTICAL APPLICATIONS: Pomelo being a rich source of bioactive nutrients faces slow commercialization due to immediate bitterness (naringin), thus in the present study, Citrus macroptera waste including peel and pomace was efficiently utilized to produce naringinase enzyme for the debittering of pomelo juice. Also, adsorption kinetics of naringin using macroporous resin is presented, which will further help in the selection of optimum time and dose for the removal of naringin from the juice. More importantly, their effect on juice composition helped in the selection of the potential candidate for the industrial purpose with negligible or minimum loss of nutrients.

Ougan juice debittering using ultrasound-aided enzymatic hydrolysis: Impacts on aroma and taste

The optimal sonication conditions (40 kHz, 80 W/L and 60 min) during Ougan juice debittering by Aspergillus niger koji extract were established. Enzymatic hydrolysis degrees of naringin and limonin were enhanced to 89.90% and 36.16%, and enzymatic hydrolysis time was shortened by 33%. Sonication significantly enhanced activities of α-l-rhamnosidases, β-glucosidases and limoninases from A. niger koji extract and facilitated break of CO bonds in naringin (p < 0.05). These accounted for the enhanced enzymatic hydrolysis degrees and velocities of bitter compounds. Meanwhile, sonication lowered 40%, 7% and 21%, 13%, 11%, 25% of bitter, sour tastes and green, citrus-like, floral, woody notes, but enhanced 18% and 15% of fruity and sweet notes, resulting in 38% and 33% increases in over-all taste and aroma scores. Lowered levels of bitter compounds, organic acids, green, citrus-like, floral, woody aroma compounds and enhanced levels of fruity, sweet aroma compounds caused by sonication accounted for the flavor improvements.

Front Cover: Covalent Immobilization of Naringinase over Two‐Dimensional 2D Zeolites and its Applications in a Continuous Process to Produce Citrus Flavonoids and for Debittering of Juices (ChemCatChem 18/2020)

Front Cover: Covalent Immobilization of Naringinase over Two‐Dimensional 2D Zeolites and its Applications in a Continuous Process to Produce Citrus Flavonoids and for Debittering of Juices (ChemCatChem 18/2020)

Covalent Immobilization of Naringinase over Two‐Dimensional 2D Zeolites and its Applications in a Continuous Process to Produce Citrus Flavonoids and for Debittering of Juices

AbstractThe crude naringinase from Penicillium decumbens and a purified naringinase with high α‐L‐rhamnosidase activity could be covalently immobilized on two‐dimensional zeolite ITQ‐2 after surface modification with glutaraldehyde. The influence of pH and temperature on the enzyme activity (in free and immobilized forms) as well as the thermal stability were determined using the specific substrate: p‐nitrophenyl‐alpha‐L‐rhamnopyranoside (Rha‐pNP). The crude and purified naringinase supported on ITQ‐2 were applied in the hydrolysis of naringin, giving the flavonoids naringenin and prunin respectively with a conversion &gt;90 % and excellent selectivity. The supported enzymes showed long term stability, being possible to perform up to 25 consecutive cycles without loss of activity, showing its high potential to produce the valuable citrus flavonoids prunin and naringenin. We have also succeeded in the application of the immobilized crude naringinase on ITQ‐2 for debittering grapefruit juices in a continuous process that was maintained operating for 300 h, with excellent results.

Isolation of naringinase producing soil bacteria from &lt;em&gt;Psidium guajava&lt;/em&gt; L. and &lt;em&gt;Terminalia chebula&lt;/em&gt; Retz and its enzymatic activity

In commercial citrus juice, bitterness causes serious challenges in fruit juice manufacturing industries. The prime cause of bitterness is due to the presence of naringin compound. It is noted that microbial enzyme retains some specific catalytic reactions of physicochemical and biological properties possessing high degree of industrial and medical applications. The microbial enzyme naringinase can be immobilized for industrial use to reduce the cost of debittering of juice. Since environment friendly industrial biocatalysts are economically more viable, so the focus on the present study is debittering of juice at low cost without using chemicals which alter the nutrient composition. In the present study, four strains of naringin degrading bacteria were isolated from the soil of Psidium guajava L. and Terminalia chebula Retz in Dibrugarh University Botanical Garden and were investigated for naringinase activity of soil microbes and their growth conditions at different parameters. The turbidity of cell culture and concentration of proteins in the culture media have been utilized for the optimization of various growth conditions, like temperature and pH for microbial growth. The optimal growth of the four isolates was observed in a media of pH 6 and selected for further study. All the four isolates showed good extracellular naringinase activity. Among the four isolates, oval and rod shaped gram positive bacteria showed the highest specific activity (405.31 U/mg) and lowest activity was shown by rod shaped gram negative bacteria (231.77 U/mg). Furthermore, rod shaped isolate exhibited maximum growth and highest protein content among the four isolates. These results suggested that in addition to the naringinase enzyme, some other proteins were also produced by the isolates. These proteins might have some significance in supporting the growth of the microorganisms.

Nano-encapsulation of naringinase produced by Trichoderma longibrachiatum ATCC18648 on thermally stable biopolymers for citrus juice debittering.

Characteristics of naringinase nano-encapsulated forms on different carrier materials (chitosan and alginate polymers) were investigated in this study. Screening of twelve fungal isolates for naringinase production indicated that Trichoderma longibrachiatum was the most promising. Grapefruit rind was used as a substrate containing naringin for naringinase production. TEM micrographs showed that chitosan nano-capsules were applied for the production of morphologically homogeneous enzymatic nano-particles with high enzyme encapsulation efficiency, small asymmetric sizes (from 15.09 to 27.07 nm with the mean of 21.8 nm) and rough surfaces compared to nano-encapsulated naringinase in alginate which showed nano-particle size (from 33.37 to 51.01 nm with the mean of 43.03 nm). It was revealed that the highest naringinase activity was found in case of chitosan nano-capsule naringinase compared to alginate nano-capsule one. Thermogram analysis (TGA) showed that the free enzyme loses about 92% of its weight at approximately 110°C, while the nano-encapsulated ones show more stability at higher temperatures. Conclusively, the nano-capsulation process improves the kinetics and operational stability so could be useful as a debittering agent for various thermal processing applications in citrus juices industries which makes the fruit juice more acceptable and cost-effective to the consumer.

Industrial orange juice debittering: effect on volatile compounds and overall quality attributes

SummaryThis study was aimed to analyse the impact of the debittering process (DP) in the overall sensory properties of orange juice (OJ) (Citrus sinensis L. Var. Salustiana). The fresh industrial squeezed orange juice (FOJ) and the corresponding OJ after the DP (DOJ) were taken. No significant differences were found in acidity, pH and total soluble solids. Colour was evaluated by image analysis (DigiEye System). Hue and lightness were lower (more reddish and darker) after debittering (P &lt; 0.001). Odour profile (limonene, α‐pinene, ethyl butanoate, octanal, linalool, citral and terpineol) decreased significantly (from 16% to 61% on average) as well as total phenolic compounds measured by Folin–Ciocalteu (P &lt; 0.05). The influence of the debittering on the perceived colour, smell and taste was evaluated by paired comparison tests. The naïve panellists found significant differences not only in taste but also in colour and aroma between FOJ and DOJ; however, preference was not clear.

Industrial orange juice debittering: Impact on bioactive compounds and nutritional value

The impact of an industrial debittering process (DP) on nutritional and bioactive compounds in orange juice (OJ) was studied. The DP was aimed at removing bitter components in OJ by physical adsorption in a resin. The levels of bioactive compounds (carotenoids, ascorbic acid and phenolics), total antioxidant activity and the colour in the fresh orange juices (non-debittered) and in the debittered counterparts were measured. The results demonstrated that the carotenoid contents were not significantly affected by the treatment. However, the debittered orange juices showed a reduction (p<0.001) of 26% in ascorbic acid, 32% in hydroxycinnamic acids, 28% of flavones and 41% of flavanones in comparison with the non-treated juices. The antioxidant activity of the hydrophilic fraction (HF) was significantly higher (p<0.05) in untreated juice than in debittered juices. Some colour parameters (L∗, a∗ and hab) were also affected. Discriminant analysis revealed that the canonical function related to the levels of HF compounds allowed a 100% correct classifications of the different types of juices.

Immobilization of naringinase from Aspergillus niger CECT 2088 in poly(vinyl alcohol) cryogels for the debittering of juices

Naringinase, induced from Aspergillus niger CECT 2088 cultures, was immobilized into a polymeric matrix consisting of poly(vinyl alcohol) (PVA) hydrogel, cryostructured in liquid nitrogen, to obtain biocatalytically active beads. The effects of matrix concentration, enzyme load and pH on immobilization efficiency were studied. Between 95% and 108% of the added naringinase was actively entrapped in PVA cryogel, depending on the conditions of immobilization used. The optimal conditions were: 8% (w/v) PVA at pH 7 and 1.6–3.7 U ml −1 of enzyme load. The pH/activity profiles revealed no change in terms of shape or optimum pH (4.5) upon immobilization of naringinase. However, the optimum temperature was shifted from 60 °C to 70 °C and the activation energy of reaction, E a, was decreased from 8.09 kJ mol −1 to 6.36 kJ mol −1 by immobilization. The entrapped naringinase could be reused through six cycles (runs of 24 h at 20 °C), retaining 36% efficacy for the hydrolysis of naringin in simulated juice.

PH influence on the consumption of limonin species by Rhodococcus fascians cells

The ability of Rhodococcus fascians cells to degrade limonin and limonin species (limonoate, limonoate-D-ring lactone and limonoate-A-ring lactone) was checked against pH. These studies showed a marked effect of pH on cell growth mainly due to substrate availability (limonin species). Evolution of limonin and its species within the medium were followed at different pH values. The best substrate for Rhodococcus fascians at pH 7.0 was limonoate whereas at pH 4.0 to 5.5 it appeared to be limonin. Results suggest that the citrus juice debittering process start only once the natural precursor of limonin (limonoate A ring lactone) has been transformed into limonin, the equilibrium displacement being governed by the citrus juice pH.

Fiber entrapment of naringinase from Penicillium sp. and application to fruit juice debittering

Naringinase from Penicillium sp. was immobilized on cellulose triacetate by the fiber entrapment method. Although the optimum pH (3.7) and optimum temperature (55°C) of the fiber-entrapped enzyme were similar to those of the native form, the immobilized enzyme had better heat stability. Kinetic studies showed that the immobilized enzyme had higher K m values than its native form. When this immobilized naringinase was successively used in a column reactor for the hydrolysis of ρ-nitrophenyl-α- l-rhamnoside or naringin in a simulated fruit juice system or grapefruit juice, the enzyme column could be operated with satisfactory stability. In addition, when the natural grapefruit juice was recycled through the column reactor, no column blocking or filtering action of the catalyst bed was observed.