A New Perspective from the Acid–Base Electronic Theory: How Do Acid Preparations Regulate Oxidative Stress?

I. On Oxidative Stress

Oxidative stress refers to the physiological phenomenon in animals characterized by elevated levels of reactive oxygen species (ROS). ROS are byproducts of normal metabolic processes and their presence is inevitable. Under normal conditions, ROS in an animal’s body exist in a dynamic equilibrium: they are continuously produced while also being steadily scavenged by the organism.

In practical animal production, farming-environment factors—such as high temperatures (heat stress), low oxygen levels (caused by excessively high stocking densities), and feed contamination (moldy feed, oxidative spoilage, excessive heavy metals, and antibiotic residues)—as well as changes in animals’ physiological states—such as weaning, pregnancy, lactation, and disease—and exogenous factors—such as bacterial, viral, and parasitic infections—can all trigger oxidative stress responses in animals. Aquatic animals face even greater oxidative stress than terrestrial animals. Consequently, increased attention to and research on oxidative stress to reduce its negative impacts on animal farming has become a hot topic in the industry.

II. Effects and Regulatory Mechanisms of Oxidative Stress

2.1 Effects of Oxidative Stress

In actual production, high morbidity rates in livestock and poultry, low survival rates in young animals, reproductive disorders, and declining quality of animal products are all associated with oxidative stress.

Figure 1. Factors causing oxidative stress and their potential effects

When animals experience oxidative stress, elevated levels of reactive oxygen species (ROS) within the body can damage vital biomolecules such as lipids, proteins, and DNA, leading to cell membrane disruption, reduced enzyme activity, and decreased RNA/DNA stability, ultimately triggering apoptosis. Oxidative stress affects animal cells, tissues, and organs almost universally, but its effects are often first evident in the intestine. The intestinal tract serves not only as the primary site for food digestion, absorption, and metabolism but also as an organ involved in the secretion of physiological fluids and immune responses. When subjected to oxidative stress, excessive ROS attack can precipitate intestinal disorders, manifesting as intestinal injury, altered intestinal mucosal permeability, and abnormal secretions from associated glands, thereby resulting in impaired intestinal function and enteric infections. Weaning stress in piglets is perhaps the most typical example of oxidative stress.

2.2 Regulatory Mechanisms of Oxidative Stress

Reactive oxygen species (ROS) are generated during normal metabolic processes in animals, so the organism itself possesses mechanisms to regulate them. Among these, Nrf2 (nuclear factor erythroid 2–related factor 2) is widely recognized as the master nuclear transcription factor regulating cellular oxidative stress. Under normal conditions, Nrf2 binds to its adaptor protein Keap1 (kelch-like ECH-associated protein 1), which keeps it inactive and sequesters it in the cytoplasm. Keap1, a multi‑domain repressor belonging to the Kelch family, acts as a sensor of oxidative stress; it typically exists as a dimer and negatively regulates the Nrf2 pathway. Together, Nrf2 and Keap1 control the expression of numerous genes encoding detoxifying enzymes and antioxidant proteins, thereby modulating intracellular ROS levels and enabling the organism to cope with oxidative stress.

Figure 2. 3D structure of the Keap1 protein

Following oxidative stress, Nrf2 is released from the cytoplasm and translocates into the nucleus, where it binds to the antioxidant response element (ARE) and activates the expression of a series of ARE‑driven genes, thereby mediating crucial cellular protective mechanisms. Nrf2 is widely conserved, ranging from lower insects to higher mammals, and is highly expressed in multiple tissues and organs—including the skin, lungs, gastrointestinal tract, liver, and kidneys—that are exposed to adverse environmental challenges.

It is worth noting that while activation of the Nrf2 protein can induce the expression of numerous cytoprotective genes, its excessive or dysregulated expression can also be detrimental to the organism. This may explain why some antioxidant agents exhibit optimal efficacy only at appropriate doses.

(Recommended reading: "nuacid Nutrition"《 How to Interpret the Signal Code Under Heat Stress Conditions in Poultry (Part 1) 、 How to Interpret the Signal Code Under Heat Stress Conditions in Poultry (Part II) 》，to learn more about the mechanisms by which oxidative stress affects animal physiology.)

III. Advances in Research on Antioxidant Additives

Animals possess their own intrinsic mechanisms; however, when oxidative stress occurs, additional antioxidants are required to help maintain the balance of the animal’s ROS-scavenging system. Such ROS scavengers are commonly referred to as antioxidants (or free-radical scavengers). Currently, antioxidants that have been extensively studied and applied can be broadly classified into two main categories: enzymatic antioxidants and non-enzymatic antioxidants.

3.1 Enzymatic Antioxidants

Enzymatic antioxidants are enzyme systems located in cell membranes and organelles that specifically neutralize free radicals. By scavenging relevant free radicals, they inhibit lipid peroxidation. The most representative examples include superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT). All of these enzymes achieve their detoxifying effect by catalyzing disproportionation reactions that convert free radicals into less reactive species.

3.2 Non-enzymatic Antioxidants

Non-enzymatic antioxidants are substances that can directly scavenge free radicals or act as cofactors for enzymatic systems, thereby enhancing their ability to neutralize free radicals. There is a large variety of non-enzymatic antioxidants, and this number continues to grow as research into antioxidant defense mechanisms advances. They mainly include the following major classes: vitamins, synthetic compounds, transition metals, and natural products.

Vitamin antioxidants mainly include: vitamin A, vitamin C, vitamin E, and β-carotene.

Synthetic antioxidants include ethoxyquin (EMQ) (EMQ, also known as ethoxyquin, is currently banned from addition), tert-butylhydroquinone (TBHQ), butylated hydroxytoluene (BHT), and butylated hydroxyanisole (BHA).

The category of trace metals includes active elements such as the trace elements selenium, copper, zinc, and manganese, with organic forms generally exhibiting superior efficacy.

Natural-product antioxidants refer to traditional Chinese medicinal herbs or plant extracts. They include a wide variety of compounds such as flavonoids, polyphenols, saponins, and polysaccharides. Due to their inherent biocompatibility, natural-product antioxidants hold the most promising prospects for future applications.

IV. Lewis Acid–Base Electronic Theory and Oxidative Stress

The Lewis acid–base theory, also known as the electron-pair theory of acids and bases or the generalized acid–base theory, is an acid–base theory proposed in 1923 by the American physical chemist Gilbert N. Lewis. It defines an acid as any molecule, group, or ion that can accept an electron pair, and a base as any molecule or group that can donate an electron pair.

Under Lewis acid–base theory, an acid can be simply described as an electron-deficient species. In biological macromolecules such as proteins, the N and O atoms in local functional groups—such as –NH2, –OH, and –C=O—each possess lone pairs of electrons in their outermost shells. These lone pairs can donate to electron-deficient Lewis acids, thereby participating in a broad sense of acid–base reactions.

Figure 3. Diagram of the binding between a Lewis acid and the active site of Keap1.

In Figure 3, a Lewis acid accepts coordinating electrons from protein residues via hydrogen bonding, p–π interactions, and other mechanisms, thereby binding to the active site of the Keap1 protein. This interaction has the potential to activate the Keap1–Nrf2 signaling pathway, suggesting that the compound may function as an enzyme cofactor-type antioxidant.

In addition to elucidating the relationship between acids and oxidative stress, Lewis acid–base theory also provides valuable guidance for evaluating the activity of antioxidants.

Currently, antioxidants that directly scavenge ROS generally achieve their antioxidant effect by undergoing chemical reactions with free radicals—either by abstracting or donating electrons to quench the radicals. Therefore, an evaluation system containing relatively stable free radicals and well-characterized reference standards can be established to assess antioxidants. Although in vitro assays are influenced by differences in biological affinity, they still provide useful reference value.

Figure 4. Computer-aided QSAR methods for predicting the structure–activity relationships of natural products.

However, currently there are no effective in vitro evaluation methods for enzyme‑assisted antioxidants. Nonetheless, based on Lewis acid–base and electron theory as well as other empirical potential functions, and with the aid of computer simulations, it is possible to perform reliable structure–activity predictions and evaluations for these compounds. Conducting research and development on animal antioxidant mechanisms under the framework of Lewis acid–base theory can more clearly elucidate the fundamental modes of action and guiding directions for antioxidant development, thereby better addressing the increasingly severe oxidative stress challenges faced by large-scale livestock production. 
 

In 2021, NIC (nuacid Company Innovation Center) pioneered the introduction of Lewis acid–base electronic theory into the agriculture and animal husbandry sector and developed a series of acidic formulation products, including TaiSuan. This product line has demonstrated significant efficacy in blocking virus transmission and mitigating viral impacts on animals. Relevant research findings have been published in international journals (see “The Effect of Monolaurin on Viral Load and Inflammatory Cytokines in PEDV-Challenged Piglets” The original journal article can be found at https://doi.org/10.3389/fvets.2022.980381; please feel free to inquire!

Written by: Liu Jian

Reviewed by: Hu Youjun

Arranged by: Li Weitao