Cold stress in plants has been extensively studied and can affect various stages of plant’s life cycle, from seed formation to development, causing damage to cell membranes, impairing cell division, and disrupting water absorption. Consequently, researchers have focused on mitigating the impacts of abiotic stress by investigating bioactive molecules and biostimulants derived from various organisms, which enhance tolerance mechanisms in plants. In aquatic environments, macro- and microalgae have emerged as key sources of plant elicitors, providing extractable molecules such as polysaccharides, polyamines, polyphenols, and amino acids that enhance plant defense responses. Similarly, certain terrestrial plants have shown potential as sources of biostimulant compounds.
Introduction
The search for new bioactive molecules as a solution to reduce agrochemicals in agriculture has led to countless experiments focusing on genetic resources, either from terrestrial or marine environments. In this context, extracts and molecules derived from plants and algae have proven effective as elicitors of plant development and health, enhancing root growth, flowering, and tolerance to adverse environmental conditions.
Particularly in marine environments, microalgae and macroalgae have been identified as potential sources of biofertilizers and biostimulants for plants. They contribute to nutrient transport, plant protection, and growth, besides serving as nutritional additives or elicitors for tolerance induction through bioactive molecules. These include polysaccharides such as ulvans, fucans, alginates, and carrageenans, as well as phytohormones, betaines, polyamines, and amino acids. Algal biomass and their constituents have been extensively studied for applications in agriculture. Algal species from all divisions, including Chlorophyta (green), Rhodophyta (red), and Phaeophyta (brown), have been used for mitigate the impacts of abiotic stress in plants. The results from foliar and soil applications have shown, for instance, increased root emission and shoot growth, along with enhanced defense responses and plant tolerance to (a)biotic stress factors.
In addition to algae, many terrestrial plant species contain bioactive molecules that may benefit cultivated plants. Plant extracts and oils can serve as additives, biofertilizers, biostimulants, biopesticides, and protectors of crop species. The market for these biomolecules has been steadily expanding, driven by farmers’ recognition of their effectiveness in low-carbon food production systems.Given the potential of algae and plant-based biostimulant compounds, research has focused on developing new technologies to enhance plant tolerance to abiotic stresses, such as water deficit, salinity, and thermal stress – common climatic events that pose significant challenges and lead to major production losses. Among these, cold stress is particularly notable for causing substantial losses, both in the field and post-harvest. However, studies addressing this issue remain limited, especially regarding its impact on global agricultural production. Thus, this review aims to present and critically discuss the findings of peer-reviewed articles and books that have described and confirmed the effectiveness of seaweed-based and plant biostimulants in mitigating cold stress in various crops.
Biostimulants – definition, market, and legislation
Biostimulants are products used in agriculture that can be applied to plants and soil and are capable of regulating and/or enhancing physiological processes in crops. Considered environmentally safe and cost-effective alternatives to traditional fertilizers and chemical treatments, they minimize negative impacts on ecosystems while promoting sustainable agricultural practices. They influence plant biochemistry through various metabolic pathways, enhancing morphological yields, increasing quality and shelf life, aiding in the absorption of essential nutrients, and boosting tolerance to abiotic stresses.
In industry, the definition of biostimulant was originally proposed in 2012, highlighting that these are plant-based products containing compounds and/or microorganisms that stimulate natural processes when applied to plants and the rhizosphere. Importantly, they do not fit into the regulatory framework for pesticides, as they do not have a direct action against pests. According to du Jardin (2015), a plant biostimulant refers to any substance or microorganism applied to plants to improve nutritional efficiency, enhance tolerance to abiotic stress, and/or improve crop quality characteristics, irrespective of its nutrient content.
Apparently, the term was coined by horticultural experts to describe compounds that promote plant growth without being defined as nutrients, soil amendments, or pesticides. However, the discussion on this topic can be traced back to 1933, when Professor Filatov in the Soviet Union introduced the concept of biogenic stimulants. Filatov suggested that biological materials from various organisms, including stressed plants, could influence metabolic and energy processes in plant, animal, and even human cells. Other contributions to the discussion came from Blagoveshchensky in 1945, when he described that biogenic stimulants, such as organic acids, would have a stimulating effect due to their dibasic properties, which can increase enzymatic activity in plants. Filatov, however, continued to expand the discussion by not limiting it to organic acids.
Building on these foundational insights, recent advancements have highlighted key benefits for crop production, such as enhanced growth, increased yield, and improved fruit quality. Additionally, there has been progress in mitigating biotic and abiotic stresses, enhancing beneficial soil microbiota and improving nitrogen use efficiency and crop yield. Other notable advancements include increased shelf life and the enhancement of bioactive compounds in plants.
In the sense of the legal framework, someone describes a discrepancy among various countries, states, and regions regarding the categorization and registration of biostimulant products, which can impact trade hinder further developments. The terminologies associated with biostimulants in legislation include planting conditioners, other fertilizers, supplements, soil correctives, plant fortifiers, and phytofortifiers, for example. In addition to the terminology used for product registration, many jurisdictions require detailed information identifying all substances present in the product, while others allow registration without complete identification, noting that these products are considered matrices of complex composition.Finally, one of the biggest obstacles to the standardization of biostimulant products regarding their production, registration, marketing, and use is the large number of molecules in their composition that are claimed to have stimulant properties for plant development, as well as the differing regulatory requirements. Because of this, different countries have sought to harmonize production and evaluation processes in the search for a common standard for these bioactive molecules.
Compounds of interest
Over the years, researchers have proposed different categorizations of biostimulant products, especially based on the main component or modes of action. In European Union countries, e.g., this information is included on the label of commercialized products and is serves as a means of registering biostimulants. The classification based on the origin of the raw material is the most commonly used. Thus, biostimulants can be divided into two groups: those of biological origin, where the components and molecules come from the plant or other organic sources, and those of physical or chemical origin.
In this context, certain groups of compounds can be mentioned, including phytohormones, polyamines, amino acids, polyphenolic compounds, and polysaccharides. Given the significance of these compounds, the literature review describes various biological activities of marine algae for use as biostimulants in plants. These activities include increased productivity and quality of agricultural crops, enhancement of biochemical compounds, improved yield and essential oil content, as well as increased tolerance to (a)biotic stress factors. Similarly, plant biostimulants can also achieve comparable results. Thus, it is evident that these substances possess considerable potential for enhancing plant growth and resilience.
Regarding phytohormones, these low-molecular-weight compounds play crucial roles in regulating plant growth and development, being associated with both morphophysiological and biochemical responses. One of the main phytohormones is abscisic acid (ABA), which has long been considered the primary phytohormone linked to stomatal regulation and plant stress responses. However, other phytohormones also play significant roles in plants, including cytokinins, auxins, gibberellins, and ethylene.
ABA, a bioactive terpenoid phytohormone, is recognized as one of the primary components in seaweed extracts. Its significance in plant physiology arises from its pivotal role in regulating various stress responses and developmental processes. As a biostimulant, ABA enhances several biochemical parameters, such as chlorophyll content, antioxidant enzyme activity, and osmotic adjustment compounds, which collectively contribute to improved plant resilience under adverse conditions.
Cytokinins, along with auxins, are responsible for complex biochemical processes within plant systems, although many of these processes remain to be fully elucidated. Together with auxins, they are well-known for their roles in plant growth and are commonly found in macroalgae, where they, along with macro and micronutrients, enhance developmental processes in plants . Additionally, the review reports its action in plant defense mechanisms against light, temperature, drought, osmotic, salinity, and nutritional stress factors, as well as a complete response to certain plant pathogens and herbivores.
Auxins constitute a group of low-molecular-weight phytohormones essential for plant growth and development. These molecules regulate multiple physiological processes, including cell division, elongation, differentiation, organogenesis, and responses to environmental stimuli. The most abundant and biologically active auxin is indole-3-acetic acid (IAA), although other naturally occurring auxins, such as indole-3-butyric acid (IBA), 4-chloroindole-3-acetic acid (4-Cl-IAA), and phenylacetic acid (PAA), also contribute to plant development. In addition to growth regulation, auxins play a key role in stress responses, particularly in modulating root architecture under adverse conditions, such as low temperatures.
Gibberellins are classified as carboxylated diterpenoids, with modifications in their chemical structure depending on their source. Their biosynthesis occurs through the methylerythritol 4-phosphate (MEP) pathway, and they are primarily found in higher plants. The most abundant and the first gibberellin to be characterized was the gibberellic acid (i.e., gibberellin A3), derived from the fungus Fusarium fujikuroi. This compound is still produced on an industrial scale for use in agriculture. Notably, among their various biochemical properties, these molecules in biostimulants contribute to hormonal regulation. Gibberellins, due to their interactions with other phytohormones, help maintain a balance in the physiological state of plants during both abiotic and biotic stress.
Ethylene, in turn, is one of the simplest and most accessible molecules for biosynthesis in plant systems. Recognized as an essential growth regulator, ethylene is involved in a variety of physiological and developmental processes, ranging from growth regulation to the induction of fruit ripening, as well as in multiple stress responses. In response to biotic stress, ethylene plays a role in disease resistance by inducing the expression of genes involved in plant defense, such as pathogen-related proteins (PRs) and phytoalexins. Moreover, others have shown that ethylene content is augmented in responses to abiotic stress factors, such as drought, flooding/hypoxia, osmotic pressure, salinity, heat, cold, and heavy metals.
In the group of plant growth regulator compounds, polyamines play an important role. These small, naturally occurring organic polycations, with contain two or more amino groups, are found in both prokaryotes and eukaryotes, and they are crucial for various physiological processes in plants, such as growth and the increase of bioactive compounds. They have also demonstrated the ability to act in stress responses. Among the main exogenous polyamines used as potential biostimulants are putrescine, spermidine, spermine, and termospermin.
Putrescine is a simple polyamine composed of two amino acids, ornithine and arginine. It plays a crucial role in cell growth, differentiation, and apoptosis. Putrescine is involved in various physiological processes, including stress responses and the regulation of gene expression in plants. In addition to its direct relation to plant growth and developmental processes, it contributes to tolerance against various abiotic stresses, such as salinity, low and high temperatures, and drought.
Spermidine is a higher-order polyamine derived from putrescine. It is involved in cellular processes such as cell proliferation, differentiation, and apoptosis. Additionally, spermidine has been linked to the regulation of ion channels and is recognized for its antioxidant properties, which contribute to stress tolerance in plants.
Spermine is another polyamine formed from spermidine. It has a more complex structure and is involved in various cellular functions, including protection against oxidative stress and regulation of ion transport. Spermine also plays a significant role in plant growth and development. Furthermore, it serves as a regulator of biosynthetic pathways, decreasing ethylene levels while increasing phenolic compounds and sugars. This modulation enables plants to better tolerate harmful abiotic effects, such as drought and salinity.
Thermospermine is a polyamine derivative of spermine, particularly important in certain plant species and associated with plant growth-related genes. Additionally, it has been linked to adaptation to environmental stresses.
Amino acids are essential for plant metabolism, serving not only as building blocks for proteins but also as key regulators of stress responses. Proline aids in osmotic adjustment and provides protection against oxidative stress, while glutamate acts as a precursor for proline and GABA (γ-aminobutyric acid), both crucial for stress adaptation. Arginine contributes to nitrogen storage and oxidative stress signaling, whereas methionine and cysteine help regulate redox homeostasis through glutathione and polyamines. Lysine catabolism produces proline and pipecolic acid, enhancing plant stress tolerance, while tryptophan is vital for auxin biosynthesis and root development. These amino acids interact with phytohormones like ABA and ethylene, influencing drought resilience and salinity responses. Advances in metabolic engineering have targeted amino acid biosynthesis to improve stress tolerance, revealing both benefits and metabolic trade-offs in crop improvement.
Polyphenolics are a class of secondary metabolites that play crucial roles in plant metabolism. They are widely distributed throughout various plant parts, including leaves, flowers, fruits, and seeds. A broad diversity of polyphenols exists, and these compounds positively affect plant growth and development while also serving as elicitors of tolerance to (a)biotic stress factors. Examples include salicylic acid, anthocyanins, gallic acid, quercetin, and the stilbene resveratrol.Finally, polysaccharides are macromolecules formed by long chains of monosaccharides, which are simple sugars. In plants, these compounds play essential structural and functional roles, serving as fundamental components of plant cell walls. The application of polysaccharides derived from both marine and terrestrial sources has shown significant positive responses in plants. The study indicates that alga-derived polysaccharides such as agar, alginate, and carrageenan can enhance seed germination and plant vigor, increase nutrient absorption from the soil, and protect plants against various abiotic and biotic stresses, including salinity, drought, temperature extremes, and pathogens. Similarly, plant-derived polysaccharides like as xyloglucan, found in various plant species, contribute to soil aggregation and the maintenance of its physical properties. Under stress conditions such as salinity, xyloglucans can induce the expression of tolerance genes, mitigating the negative effects of oxidative stress in plants.
Adverse weather conditions and impact on agriculture
Global warming, industrialization, and climate change are interconnected through complex relationships. Significant increases in greenhouse gas (GHG) emissions, mainly carbon dioxide (CO2), from the burning of fossil fuels, have occurred since the beginning of industrialization. The increase in CO2 concentrations and other GHGs in the atmosphere traps heat and contributes to global warming, altering climate events such as droughts, more frequent and intense heat waves, changes in precipitation patterns and temperatures, and the melting of glaciers, as well as rising temperatures and sea level.
Such changes can directly impact crop production due to the adverse environmental conditions which plants will be exposed.Climate plays a crucial role in determining the success or failure of agricultural practices and overall crop productivity. Climatic conditions such as temperature, sunlight, and rain directly influence the growth and development of plants, as each crop has specific temperature and photoperiod requirements for optimal growth. These variables affect the progression of crop stages across a wide spectrum, encompassing both vegetative and reproductive phases. Deviations in climate patterns can disrupt these phenological stages. Furthermore, precipitation impacts water availability by altering soil moisture levels, with insufficient or excessive precipitation leading to droughts or floods, respectively, which consequently influences the availability of nutrients that plants can absorb. According to the FAO, extreme adverse weather events such as droughts, floods, heat waves, and cold spells significantly impact agriculture and affect crop yields. In summary, climate change threatens global crop production, requiring adaptive agronomic strategies to enhance resilience. Among these strategies, biostimulants play a key role in mitigating stress by modulating plant physiology and improving tolerance to environmental challenges.
Cold stress
Temperature plays a fundamental role in the growth and development of plants, and extremely low or high temperatures cause great stress, restricting natural processes at the genetic, biochemical, and physiological levels. Especially in recent decades, due to climate change this environmental factor has been widely discussed by experts in the field of food production, who seek to explore new possibilities to face this challenge. In fact, low temperatures have represented one of the main environmental factors for the suppression of agricultural crops, which leads to drastic losses annually in the world.
Estimates indicate losses of 50% in the field due to plant stresses resulting from temperature changes. In 1972 and 1976, losses of 42% and 37%, respectively, were reported in rice production in the northeast region of China due to severe cold stress. In horticulture, for example, countries such as France, Germany, Italy, Belgium, Switzerland, and the USA recorded large agricultural losses due to increased frosts in the past years.
Regarding the low temperatures, plants can be categorized as cold-sensitive, cold-tolerant, and freeze tolerant. Plants sensitive to cold are those that die at the beginning of stress conditions, not resisting extreme conditions. In turn, freezing-tolerant plants are those that face the greatest cold stress. Due to their cold adaptation characteristics, the geographic distribution of plants is directly affected, where tropical and subtropical plants (e.g., rice, soybeans, tomatoes, corn, and cotton) are sensitive to cold stress, lacking the ability to acclimate to cold, as temperate species (e.g., barley, wheat, and rye) have a greater capacity for freezing tolerance.
Low temperatures above freezing conditions can slow plant metabolism, decreasing photosynthetic levels, leaf growth, and early senescence. In conditions lower than freezing, the development of buds can be seriously impaired, resulting in the destruction of rehydrated buds. In cases of frost, plant tissues dehydrate, resulting in an increase in the concentrations of osmolytes in the cell cytoplasm, consequently, resulting in the rupture of the plasma membrane. Generally speaking, damage to plant tissues can lead to poor growth, delayed flowering, reduced fruit set, and lower productivity overall. The severity and duration of cold stress can determine the extent of yield and quality losses and, in extreme cases, result in plant death or total crop loss, which can extend beyond the immediate growing season and affect the production of future crop cycles.
Plants respond to cold stress through several physiological mechanisms to deal with adverse conditions, namely changes in gene expression, cellular metabolism, and adjustments in water and nutrient absorption. The plant’s response can occur through different metabolic pathways, each of which can represent tolerance, attempts to avoid, escape, and recover. For example, as a response to cold stress, several pathways can be affected, such as calcium channels (Ca2+). From the moment extracellular ice is formed, which leads to water loss in cells, the integrity of cell membrane systems is affected, leading to the opening of Ca2+ channels, due to the loss of membrane fluidity. The change in Ca2+ concentration induces the effect of signaling cascades through mitogen-active protein kinases (MAPKs), which can reach transcriptomic levels. In the carbohydrate pathway, in turn, immediate reprogramming occurs, as it is necessary to avoid any type of imbalance that could cause cell damage or death. Thus, the effects of cold stress include physiological adaptations to water deficiency, such as the accumulation of various osmolytes and antioxidants, changes in phospholipid composition, production of reactive oxygen and nitrogen species, and the activation of phosphoprotein cascades. In fact, the knowledge about stress detection and plant response is a key issue so that preventive measures can be implemented at these times.In this sense, biostimulant elicitors can play a crucial role in increasing plant resilience and improving tolerance to cold stress, as they can activate the plant’s natural defense mechanisms against stresses, stimulating them to grow. Biostimulants can help in the production of protective compounds, such as antioxidants, osmoprotectors, and heat shock proteins, which help the plant to cope with the cold, and modulate gene expression in plants, activating specific genes associated with cold tolerance, increasing the plant’s ability to withstand low temperatures and reduces the negative impact of cold stress. Furthermore, biostimulants can improve various physiological processes in plants, including photosynthesis, respiration, and nutrient metabolism, which improve overall health and vigor, making them more resistant to cold stress.
Mechanisms of action of biostimulants and biofertilizers
Biostimulants enhance plant tolerance to cold stress by activating multiple protective pathways . These compounds operate through three primary mechanisms: enhancing antioxidant defense systems, accumulating osmoprotectants, and modulating hormonal signaling (Figure 1). Together, these mechanisms maintain membrane integrity, stabilize cellular proteins, and sustain essential metabolic functions, thereby mitigating the detrimental effects of cold temperatures on plant physiology.
Antioxidant defense enhancement
Reactive oxygen species (ROS) are byproducts of cellular metabolism that can accumulate under stress conditions, such as extreme temperatures. While low levels of ROS function as signaling molecules, excessive accumulation leads to oxidative damage to lipids, proteins, and DNA. Plants possess sophisticated antioxidant defense systems to maintain ROS homeostasis, and the redox-regulatory network is critical for cold stress acclimation.
Biostimulants significantly enhance these natural defense mechanisms by increasing the activities of antioxidant enzymes, including superoxide dismutase, catalase, and ascorbate peroxidase, which are crucial for scavenging ROS. Someone demonstrated that algae-based biostimulants improved the physiological condition of zucchini plants under cold stress, contributing to the limitation ROS accumulation while simultaneously providing osmoprotection.
Osmoprotectant accumulation
Osmoprotection is a critical mechanism through which biostimulants enhance plant tolerance to cold stress. Under low-temperature conditions, plants face cellular dehydration and potential ice crystal formation, which can damage cellular structures. Biostimulants stimulate the biosynthesis and accumulation of key osmolytes, including secondary metabolites (proline, betaine, and putrescine) and compatible solutes (sucrose, glucose, raffinose, fructose, and trehalose).
This elevated concentration of osmolytes within the cytoplasm maintains a lower cellular water potential than the external environment, even under cold-induced dehydration. The resulting osmotic adjustment promotes water influx into cells, preserving cellular hydration and turgor pressure – both essential for cell expansion, stomatal regulation, and overall metabolic function. Additionally, these osmolytes interact directly with proteins and membranes, preventing denaturation and maintaining the functional integrity of essential cellular components, including metabolic enzymes, photosynthetic complexes, and membrane-bound transporters.
Hormonal signaling modulation
Biostimulants exert significant effects on plant hormone pathways that regulate cold stress responses. Cold stress directly impacts auxin transport by inhibiting the recycling of transmembrane proteins responsible for hormone redistribution, which compromises the establishment of auxin gradients essential for regulated growth. While the relationship between exogenous auxin application and cold tolerance remains incompletely understood, research suggests that auxins mediate plant responses to low-temperature stress. For instance, IAA stimulates arbuscular mycorrhizal development, potentially contributing to plant adaptation under adverse environmental conditions such as cold.
Someone demonstrated that disruptions in auxin transport and signaling pathways alter root morphology under cold stress, characterized by reduced auxin levels in root tips. This reduction inhibits positive cell cycle regulators while enhancing negative regulators. Notably, exogenous auxin application promotes root growth under cold stress conditions. However, it does not fully restore normal growth, suggesting the involvement of additional factors in the comprehensive cold stress response.
Cytokinins (CKs) represent another crucial hormone class modulated by biostimulants. It is investigated the impact of CK levels on cold stress responses using Arabidopsis thaliana transformants with either enhanced biosynthesis (DEX: IPT) or increased degradation (DEX: CKX) of Cks. Under various light conditions, plants with elevated CK and auxin levels, along with increased salicylic acid concentrations, demonstrated enhanced stress acclimation. Conversely, plants with reduced CK and auxin contents displayed weakened stress tolerance. This study highlighted the critical interplay between CKs and light signaling in regulating cold stress responses. Biostimulant application stimulates cytokinin modulation, effectively improving plant responses to the physiological stress caused by low temperatures.
Ethylene, another plant hormone implicated in stress responses, exhibits context-dependent effects on cold stress tolerance. While it can contribute to acclimation, excessive ethylene accumulation can intensify cellular damage under cold conditions. Biostimulants help maintain optimal ethylene levels, preventing the potentially detrimental effects of low temperatures on cell physiology.
Conclusions, gaps and perspectives
Rising environmental challenges, notably extreme temperatures, demand innovative solutions for crop production and protection. Low temperatures pose a significant threat, leading to substantial global agricultural losses. Plants respond to cold stress through various physiological mechanisms, including changes in gene expression, adjustments in cellular metabolism, and alterations in water and nutrient absorption. Therefore, finding effective solutions to address this issue is urgent.
Bioactive elicitors from macro- and microalgae enhance plant resilience to cold by activating natural defense mechanisms, promoting the expression of cold tolerance genes, and improving physiological processes. Commercial products derived from macroalgae, particularly Ascophyllum nodosum, are among the most widely used and show promise in boosting cold tolerance by increasing gene expression, reducing chlorophyll damage, and stimulating plant growth. Similarly, plant-derived biostimulants, such as extracts and soil microorganisms, can promote resilience in crops, by enhancing nutrient uptake, improving overall plant health, and stimulating natural defense mechanisms.
There is still a need for improved guidelines and analysis parameters for determining whether a product qualifies as a biostimulant, whether it derives from a terrestrial or marine organism, or from purified molecules. Additionally, it is crucial to ascertain the most effective concentrations, optimal timing, and suitable delivery methods, taking into account variations among plant species and stress conditions. An essential aspect of research involves understanding the kinetics of absorption and distribution of certain molecules within plants.
If you are interested in biostimulants, see more details of Dora Amino Acids and Dora Seaweed Extracts.