Naringinase is an enzyme complex that can hydrolyze naringin, a bitter flavonoid found in citrus fruits, into naringenin, a non-bitter flavonoid with various health benefits. Naringinase has two glycosidase activities: α-L-rhamnosidase and β-D-glucosidase, which can cleave the rhamnose and glucose residues from naringin, respectively. Naringinase is widely distributed in nature and has been reported in fungi, bacteria, and yeasts. However, different sources of naringinase have different enzymatic properties and applications.

Naringinase has many potential applications in food and pharmaceutical industries, such as debittering of citrus juices, enhancing the aroma of wines, producing rhamnose and prunin, improving the clarity of canned oranges, and synthesizing various chiral compounds. In this blog post, we will introduce some of the recent advances and challenges in the production, characterization, and utilization of naringinase.

Production of naringinase

Naringinase can be produced by various microorganisms, such as Aspergillus niger, Aspergillus oryzae, Penicillium, Xanthomonas campestris, and Rhodotorula rubra. Among them, fungi are the main sources of naringinase, as they can produce high yields and stable enzymes. However, fungal naringinase usually requires expensive substrates, such as citrus peel or naringin, for induction and expression. Moreover, fungal naringinase is often contaminated with other enzymes, such as pectinase and cellulase, which may affect its quality and activity.

Bacterial naringinase, on the other hand, has some advantages over fungal naringinase, such as lower production cost, higher specificity, and easier purification. However, bacterial naringinase is less studied and reported, and its production is often limited by low expression levels, poor secretion, and low stability. Therefore, more research is needed to optimize the production of bacterial naringinase, such as screening for novel strains, engineering the expression systems, and improving the fermentation conditions.

Characterization of naringinase

Naringinase is a heterodimeric enzyme, composed of a heavy chain and a light chain, which are linked by a disulfide bond. The heavy chain contains the α-L-rhamnosidase activity, while the light chain contains the β-D-glucosidase activity. Naringinase belongs to the glycoside hydrolase family 78, which is characterized by the presence of a serine residue in the active site, which can form a covalent bond with the glycosidic bond of naringin.

Naringinase has a broad substrate specificity, as it can hydrolyze various glycosides containing terminal α-L-rhamnose and β-D-glucose, such as rutin, quercitrin, hesperidin, diosmin, and ter-phenyl glycosides. Naringinase also has a high affinity and catalytic efficiency for naringin, which is its natural substrate. Naringinase has an optimal pH of 8.0 and an optimal temperature of 37°C. Its activity is influenced by various factors, such as salt concentration, metal ions, and inhibitors. Naringinase is activated by calcium and magnesium ions, but inhibited by copper and zinc ions. Naringinase is also inhibited by some natural or synthetic compounds, such as naringin, naringinase inhibitor, sulfonamides, and phenylboronic acid.

Applications of naringinase

Naringinase has found wide applications in food and pharmaceutical industries, as it can hydrolyze naringin into naringenin, rhamnose, and glucose, which have various benefits and uses.

•  Debittering of citrus juices: Naringin is one of the main bitter compounds in citrus fruits, and it has a bitterness threshold of about 20 ppm in water and 30 ppm in juice. Naringin is mainly distributed in the peel, pulp, and seeds of citrus fruits, and its content varies with the origin, variety, and maturity of the fruits. Naringinase can specifically hydrolyze naringin into naringenin, which has no bitterness and has antioxidant, anti-inflammatory, and anti-cancer properties. Naringinase can also hydrolyze other bitter glycosides in citrus juices, such as neohesperidin and limonin. Therefore, naringinase can improve the quality and flavor of citrus juices by removing the bitterness.

•  Enhancing the aroma of wines: Monoterpenes are compounds that can improve the aroma of wines. They include volatile compounds and non-volatile precursors with glycosidic bonds. The non-volatile precursors consist of disaccharides (α-L-rhamnosyl-β-D-glucosides), which can be hydrolyzed by naringinase into monoterpenes and sugars. The monoterpenes can then volatilize and contribute to the aroma of wines. Naringinase can also increase the content of free naringenin in wines, which can enhance the color and stability of wines.

•  Producing rhamnose: Naringin can be hydrolyzed by the α-L-rhamnosidase activity of naringinase to produce rhamnose, which is a chiral intermediate that has important applications in organic synthesis and plant protection. Rhamnose can be used to measure the intestinal permeability, to act as a sweetener, and to produce flavoring agents.

•  Producing prunin: Prunin is a bioactive compound that has different antiviral and anti-inflammatory activities against DNA/RNA viruses. It can also act as a sweetener for diabetic patients, and as a raw material for synthesizing various drugs, such as those for expanding capillaries. Prunin can be well absorbed by the human body. Naringinase can replace the chemical to produce prunin with high purity.

•  Improving the clarity of canned oranges: The turbidity in canned oranges is characterized by the appearance of turbidity, and sometimes precipitation, in the juice, and white spots on the back of the orange segments. These white turbid precipitates are mainly composed of naringenoside, which accounts for about 57% of the turbid substances, followed by pectin and a small amount of protein. Naringenoside and pectin are most abundant in the peel and pulp of oranges, followed by the inner and outer skins and the sandbags, and less in the juice. Naringenoside is the source of bitterness in citrus fruits, and its content varies with the variety and maturity of the fruits. According to the research, the solubility of naringenoside in the canned sugar solution varies with the storage time, and is about 12-20 mg. When the content in the solution exceeds this value, crystals will precipitate, thereby reducing the transparency of the sugar solution. Naringinase can solve this problem by hydrolyzing naringenoside.

As a dedicated enzyme manufacturer and supplier, Sunson Industry Group Co., Ltd has developed wide range of enzyme products to address concerns in various industries, such as food, feed and technical industries. Naringinase and its components, α-L-rhamnosidase and β-D-glucosidase are all available and can be adapted to different situations.