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In recent decades, antioxidants have garnered significant attention for their health-promoting properties in human nutrition. However, the presence of antioxidants in plants is not merely coincidental nor primarily for human benefit. Instead, plants have evolved sophisticated biochemical mechanisms involving antioxidants primarily for their own survival, functioning as vital defenses against environmental stressors.

This article explores the intricate biochemical relationships between plant stress responses, reactive oxygen species (ROS), and the antioxidant defense systems that plants utilize to sustain their cellular integrity and health.

Plant Stress and Reactive Oxygen Species (ROS) Production

Plants encounter numerous environmental stresses, including intense light, ultraviolet (UV) radiation, temperature extremes (heat or cold), drought, salinity, and chemical herbicides. Exposure to these stressors significantly increases the generation of reactive oxygen species (ROS)—highly reactive molecules derived from oxygen metabolism.

Many individuals are familiar with the concept of harmful free radicals—molecules that possess an unpaired electron, making them highly unstable and reactive. Reactive oxygen species (ROS) are a subset (or type) of free radicals that contain oxygen and accept electrons from other molecules, making them primarily act as oxidizing agents, leading to oxidative damage in living cells.

Major Reactive Oxygen Species (ROS) in Plants

The normal metabolic processes of respiration and photosynthesis that occur within plant cells produce small quantities of highly reactive, oxygen-containing molecules—collectively called reactive oxygen species (ROS)—which are toxic by-products of these otherwise vital pathways.

The primary reactive oxygen species (ROS) produced in plant cells include:

• Superoxide anion (•O₂⁻): a negatively charged molecule produced in chloroplasts, mitochondria, and peroxisomes within plant cells, which can react readily with other molecules to form additional ROS.
• Hydrogen peroxide (H₂O₂): a less-reactive non-radical ROS that is produced in the chloroplast, mitochondria, and peroxisomes of plant cells. Its production is increased in response to biotic and abiotic stresses, allowing it to act as a diffusible signalling molecule while still posing a risk of oxidative damage when accumulated.
• Singlet oxygen (¹O₂): a highly reactive ROS that is produced during photosynthesis when chlorophyll molecules in the chloroplasts absorb excess light energy.
• Hydroxyl radical (•OH): the most reactive and potent oxidant generated in plant cells, extremely reactive and damaging, causing significant damage to cellular components, such as lipids, proteins, and DNA.
• Peroxyl radical (ROO•): a reactive ROS generated during lipid peroxidation—a harmful, free-radical-initiated chain reaction in which specific lipids (polyunsaturated fatty acids) in cell membranes are sequentially oxidised. This ROS propagates (spreads) this oxidative damage by removing hydrogen atoms from nearby lipids, converting them into new lipid radicals. This allows the reaction to continue across the membrane, leading to further oxidative damage to cellular structures. Note here that the action of other ROS, especially hydroxyl radicals (•OH), initiate lipid peroxidation. This triggers the formation of peroxyl radicals (ROO•), which then propagate the oxidative chain reaction across membrane lipids.

At lower concentrations, these reactive oxygen species (ROS) are necessary for various cellular signaling and developmental processes in plants. They function as key signaling molecules that enable cells to rapidly respond to different stimuli. ROS play a role in the control and regulation of biological processes such as growth, the cell cycle, programmed cell death, hormone signaling, biotic and abiotic stress reactions and development. The way they function in the critical role of plant stress response is by altering the expression of defensive genes which stimulate the plant’s defense mechanisms in order to induces tolerance to extreme environmental conditions and increase plant resilience overall.

However, at higher concentrations, elevated reactive oxygen species (ROS) levels cause abnormal cell signaling, cell damage through oxidative stress, and oxidative cell death.

Reactive oxygen species (ROS) are highly reactive chemically, and are known to damage DNA, lipids, carbohydrates, and proteins, the building blocks of living cells. They cause damage to plant cell organelles and membrane components (internal cell structures) and interrupt various metabolic pathways (series of linked biochemical reactions occurring within a cell) until oxidative cell death occurs.

Without effective antioxidant defense mechanisms, extensive cell damage accumulates, potentially leading to the decline and eventual death of the plant.

Plant Antioxidant Defense Systems

To counteract the harmful effects of ROS, plants have evolved complex antioxidant defense systems, categorized broadly into enzymatic and non-enzymatic antioxidants.

Antioxidants are substances that prevent or minimize oxidative damage by neutralizing reactive oxygen species (ROS). These systems work in complementary ways to maintain cellular redox balance. This balance, known as ROS homeostasis, is achieved when ROS production is matched by ROS scavenging through antioxidant compounds and enzymes.

Enzymes are specialized proteins that catalyze (speed up) biochemical reactions without being consumed in the process. In the context of antioxidant defense, enzymatic antioxidants catalytically convert harmful ROS into less reactive or harmless molecules. This distinguishes them from non-enzymatic antioxidants, which act by directly donating electrons to neutralize ROS but are often used up in the process.

1. Non-Enzymatic Antioxidant System

Non-enzymatic antioxidants include:

• Water-soluble compounds: Ascorbic acid (Vitamin C), glutathione, phenolic compounds (including flavonoids), and uric acid.
• Lipid-soluble compounds: α-Tocopherol (Vitamin E) and carotenoids.

These antioxidants function primarily by donating electrons to ROS, neutralizing them and converting them into less harmful molecules.

2. Enzymatic Antioxidant System

Key antioxidant enzymes include:

• Superoxide dismutase (SOD): Converts superoxide radicals into hydrogen peroxide.
• Catalase (CAT) and Glutathione peroxidase (GPX): Break down hydrogen peroxide into water and oxygen, preventing further ROS formation.
• Ascorbate peroxidase (APX), Peroxidase (POD), Glutathione reductase, Dehydroascorbate reductase, and Guaiacol peroxidase (GPOD): Facilitate the detoxification of hydrogen peroxide and other ROS.

Antioxidant Defense Levels

The antioxidant defenses of plants operate at multiple, sequential yet overlapping levels, involving both antioxidant molecules and enzymes. These levels include radical prevention, radical scavenging, repair of oxidative damage, and adaptive responses to oxidative stress.

Each of these defense levels plays a specific role in mitigating oxidative damage, and together they form a layered, multi-tiered response system, as outlined below:

The antioxidant defenses of plants can be described as operating at multiple sequential levels:

1. First-line Defense (Preventive Antioxidants):
   • Rapidly neutralize initial ROS or prevent their formation.
   • Involves enzymes like superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX).
2. Second-line Defense (Scavenging Antioxidants):
   • Neutralize ROS through direct scavenging by donating electrons, thus halting radical propagation.
   • Include hydrophilic antioxidants (ascorbic acid, glutathione) and lipophilic antioxidants (α-tocopherol, ubiquinol).
3. Third-line Defense (Repair and Clean-Up Antioxidants):
   • Activated after oxidative damage occurs, involving repair enzymes that restore damaged DNA, proteins, and lipids.
   • Enzymes involved include DNA polymerases, glycosylases, nucleases, proteolytic enzymes (proteases, proteinases, peptidases).
4. Fourth-line Defense (Adaptive Antioxidants):
   • Involve adaptive mechanisms utilizing signals from free radicals to trigger production and targeted deployment of antioxidants.
   • Essential for enhancing plant resilience under prolonged stress conditions.

Interaction with Beneficial Soil Microorganisms

Arbuscular mycorrhizal (AM) fungi form mutualistic associations with the roots of most terrestrial plants, facilitating improved nutrient and water uptake. In addition to these well-known benefits, AM fungi play a key role in enhancing the plant’s antioxidant defense system. This is achieved by stimulating the expression and activity of antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT), and various peroxidases, which help to neutralize reactive oxygen species (ROS) produced under stress.

This symbiosis not only improves the plant’s physiological tolerance to abiotic stresses—such as drought, salinity, and heavy metal toxicity—but also mitigates ROS-induced cellular damage, particularly in root and shoot tissues. The presence of AM fungi often leads to reduced lipid peroxidation and stabilized cellular membranes, demonstrating a direct role in moderating oxidative stress. These effects reflect a highly integrated response, in which the plant’s endogenous (internal) antioxidant system is reinforced by (external) microbial interaction, highlighting the ecological and functional interdependence between plants and soil microorganisms in stress adaptation.

Mechanisms of Antioxidant Action

Antioxidants mitigate oxidative stress by:

• Suppressing ROS formation, either by inhibiting ROS-producing enzymes or by chelating (binding) trace elements necessary for radical formation.
  This prevents the initial generation of reactive oxygen species. For example, chelating agents bind trace element redox-active metals like Fe²⁺ and Cu⁺, which would otherwise catalyze reactions (such as the Fenton reaction) that produce damaging radicals like hydroxyl (•OH). In addition, some antioxidants inhibit enzymes such as NADPH oxidases that actively generate ROS under stress conditions.
• Directly scavenging and neutralizing ROS.
  Antioxidants such as ascorbic acid, glutathione, carotenoids, and tocopherols act as electron donors, reacting with and neutralizing ROS—including superoxide anion (O₂•⁻), hydrogen peroxide (H₂O₂), singlet oxygen (¹O₂), and hydroxyl radicals (•OH)—before they can damage lipids, proteins, or DNA.
• Up-regulating the plant’s endogenous antioxidant defense responses.
  Some antioxidants and low ROS levels function as signals that activate stress-responsive genes. These genes code for antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and various peroxidases, enhancing the plant’s ability to respond adaptively to oxidative stress.

Key antioxidant molecules such as ascorbic acid and glutathione, linked through the ascorbate–glutathione cycle, function as vital redox buffers. This cycle plays a central role in detoxifying the hydrogen peroxide ROS in chloroplasts, mitochondria, and the cytosol. Ascorbate reduces hydrogen peroxide (H₂O₂) to water (H₂O) via ascorbate peroxidase, and is regenerated through a series of enzymatic steps involving glutathione. This continuous cycle maintains cellular redox homeostasis (a balance of oxidation and reduction reactions within a cell, which is crucial for maintaining proper cellular function and health), allowing plants to modulate ROS levels dynamically and support both stress tolerance and signalling.

In conclusion, plants employ sophisticated antioxidant defense systems as critical adaptations to survive and thrive amidst environmental stress. Understanding these biochemical mechanisms underscores the importance of antioxidants beyond their human health benefits, highlighting their fundamental role in plant resilience and survival strategies.

References

• Godoy, F., Olivos-Hernández, K., Stange, C., & Handford, M. (2021). Abiotic Stress in Crop Species: Improving Tolerance by Applying Plant Metabolites. Plants, 10(2), 186. https://doi.org/10.3390/plants10020186
• Pandey, Neha & Pandey, Shashi. (2014). Biochemical Activity and Therapeutic Role of Antioxidants in Plants and Humans. 10.1079/9781780642666.0191.
• Malik, & Bilal, Tanveer & Tahir, Inayatullah & Rehman, Reiaz & Hakeem, Khalid & Abdin, M.. (2013). Plant Signaling: Response to Reactive Oxygen Species. 10.1007/978-81-322-1542-4_1.