Enzymes are widely used as biocatalysts in various reaction systems in in in vitro diagnostics, biological research, and some industrial enzyme catalysis processes. Whether it is routine biochemical tests such as blood glucose, uric acid, ALT, or molecular diagnostic techniques such as ELISA and PCR, the catalytic efficiency of enzymes directly affects the accuracy and repeatability of the detection results. However, the activity of enzymes is not constant, but is significantly influenced by environmental conditions, with pH being one of the key factors. In order to maintain the acid-base balance of the reaction system, buffering agents become an indispensable component. It indirectly affects the conformation, substrate binding ability, and catalytic rate of enzymes by stabilizing the pH environment, thereby determining the success or failure of the entire reaction.

enzyme preparation

The relationship between enzyme activity and pH

Each enzyme has its specific optimal pH range, within which the enzyme has a higher catalytic efficiency. When the environmental pH deviates from this range, the charge state and spatial structure of enzyme molecules may change, resulting in impaired function of the active center and decreased catalytic ability. For example, lactate dehydrogenase (LDH) has higher activity around pH 7.4, while alkaline phosphatase (ALP) performs better under alkaline conditions at pH 9.0-10.5. If the pH of the reaction system fluctuates greatly, the enzyme may be partially inactivated, affecting the linearity of the reaction kinetics curve and leading to low detection results or poor repeatability.

How buffering agents affect enzyme activity

The main function of buffering agents is to resist the interference of external acid-base or reaction products on the pH of the system. In enzymatic reactions, certain reactions themselves can alter the acid-base state of the system. For example, glucose oxidase may produce acidic byproducts during the catalytic process, while urease breaks down urea and releases alkaline substances. If there is no effective buffering system, these changes will lead to continuous pH shift, ultimately affecting the stability and reaction rate of the enzyme.

When selecting a suitable buffer, it is necessary to consider whether its pKa value is close to the required pH for the reaction, as well as its stability under temperature changes and dilution conditions. In addition, buffering agents should not form precipitates with metal ions (such as phosphates and Ca ² ⁺), nor interfere with detection signals (such as Tris absorption in the ultraviolet region), nor should they inhibit enzyme activity.

The consequences of improper buffer selection

If the applicable pH range of the buffer does not match the optimal pH of the enzyme, even if the initial pH is correct, it may not be possible to maintain stability during the reaction process, resulting in some adverse consequences: 1. Decreased reaction rate and sensitivity; 2. The standard curve is non-linear and the quantification is inaccurate; 3. The performance differences between different batches of reagents increase; During storage, the activity of the reagent decreases and the shelf life is shortened. Therefore, the choice of buffer not only affects the immediate performance of the reaction, but also relates to the long-term stability and inter batch consistency of the reagent.

Biological buffer

Hubei Xindesheng: Providing stable and reliable buffer products

Hubei Xindesheng Material Technology Co., Ltd. focuses on the research and production of biological buffering agents. Its main products include Tris, HEPES, MOPS, CAPS, PBS, etc., which are widely used in in vitro diagnostic reagents, scientific research experiments, and industrial fields. The company adopts synthesis and purification processes to ensure product purity of ≥ 99%, and controls the content of impurities such as moisture and heavy metals. Each batch of products undergoes pH buffering performance testing to ensure stability between batches. The product provides technical documentation, supports customized packaging and bulk supply, and meets the needs of different customers.