Bromelain

Bromelain belongs to thiol proteases, and is a proteolytic enzyme extracted from pineapple, mainly found in the stems and fruits. Bromelain can be divided into stem bromelain (EC 3.4.22.4 or EC 3.4.4.24) and fruit bromelain (EC 3.4.22.5) depending on the extraction site. Because bromelain has high biocatalytic activity, it has been widely used in various industrial fields with the continuous improvement of enzyme purification technology. In bio-chemicals, bromelain can be used in the production of cheese, gelatin, and hydrolyzed proteins. In the food industry, bromelain can be used as a food additive to tenderize meat and clarify beer. In the pharmaceutical industry, bromelain is used to treat edema and various inflammations because it can dissolve fibrin and blood clots. Bromelain is harmless to normal tissues and does not affect skin grafting, so it is suitable for the treatment of small areas of deep burns.

Molecular Structure

Bromelain is an enzyme system composed of a variety of enzymes of different molecular weights and molecular structures, including at least 5 proteolytic enzymes. In addition, some bromelain also have phosphatase, peroxide, cellulase, glycosidase and non-protein substances. Bromelain is a glycoprotein with an oligosaccharide in its molecular structure consisting of xylose, fucose, mannose and N-aceltylglucosamine. Ishihara and Yasuda et al. inferred the structure of the oligosaccharide molecule of bromelain, respectively.

Figure 1. The molecular structure of the oligosaccharide molecule of bromelain speculated by Ishihara and Yasuda et al.

Properties

The isoelectric point of bromelain is 9.55, and the maximum absorption wavelength of the enzyme solution is 280 nm. Because of its composition of multiple enzyme molecules, the catalytic substrate of bromelain is diverse. It can decompose proteins, peptides, esters and amides, and its proteolytic activity is more than 10 times higher than that of papain. The essential group for bromelain to catalyze is not the oligosaccharide molecule, but the sulfhydryl group in the peptide chain. In addition, the carboxyl group and the hydroxyl group in the molecular structure of bromelain are also independent of the enzyme activity. The catalytic activity of bromelain is susceptible to pH, temperature, metal ions, EDTA, and reducing agents. The optimum pH of the enzyme is 7.1, the most stable pH range is 3.9-4.2, and the optimum reaction temperature is 55°C. Mg²⁺ and Ca²⁺ have an inhibitory effect on bromelain activity at high concentrations, but have a promoting effect at low concentrations. EDTA can chelate the metal ions required for the bromelain reaction, reducing the activity of bromelain. The hydrochloride salt of cysteine promotes the enzyme reaction rate within a certain concentration range.

Stability

Bromelain is a multi-enzyme system. Because of the different stability of different enzymes, the adaptability of bromelain to the changes of the external environment is very narrow, which undoubtedly imposes great restrictions on its industrial production. There are many factors that affect the stability of bromelain, such as temperature and humidity. Bromelain can be inactivated at high temperature and low temperature. The loss rate of enzyme activity at 4°C for 6 months is 5%-10%, and the loss rate of enzyme activity at normal temperature is as high as 10%-15%. Under high temperature conditions, a large amount of water in the air makes the flexibility of the structure of bromelain increase, and the self-hydrolysis ability is enhanced, thereby accelerating the denaturation process. Some metal ions, organic carboxylates, monosaccharides, etc. can stabilize bromelain to some extent, and this effect is achieved by stabilizing its three-dimensional conformation. In addition, the unmodified polyethylene glycol does not bind to bromelain and prevents the binding of bromelain to the catalytic substrate in the reaction system, thereby reducing its enzyme activity. However, when the succinic anhydride-activated polyethylene glycol chemically modified bromelain, the stability of the modified enzyme was greatly improved compared to the native bromelain.

Applications

• Application of bromelain in medical field

Bromelain has a good effect in the prevention and treatment of diarrhea in monogastric animals. Bromelain uses its proteolytic function to inhibit the receptor activity of pathogenic strains in the intestinal tract of animals, thereby reducing their adhesion in vivo. In most cases, bromelain is used as a drug in a certain ratio to other drugs to achieve the best clinical results. Chlorpheniramine is a class of anti-allergic drugs that, when combined with bromelain, provide excellent anti-inflammatory effects and prevent increased capillary permeability due to histamine. For male chronic prostatitis, the treatment with antibiotics plus bromelain enteric-coated tablets is significantly better than antibiotics alone.

• Application of bromelain in food industry

Bromelain is a thiol protease that directly interacts with myofibrillar proteins, hydrolyzes fibrin and connective tissue, degrades proteins into small molecules of peptides and amino acids, breaks muscle filaments and tendons, and makes meat tender and smooth. The use of bromelain to hydrolyze collagen to produce gelatin can thicken and emulsify food, and increase the content of food which is easily digested and absorbed, and has a therapeutic effect.