The first is based on the possibility that the bio-stimulant contains within it, previously unrecognized molecules that are the sole and discrete cause of the observed improvement in plant productivity. This concept emphasizes both the need for clear demonstration of plant productivity benefits and the unknown nature of the mode of action. Thus, a bio-stimulant could be defined as “a formulated product that improves plant productivity by a mechanism of action that is not the sole consequence of the presence of known essential plant nutrients, plant hormones, plant growth regulators or plant protective compounds.” By this definition, once the primary biological mechanism of bio-stimulant function has been identified it should henceforth, be subject to classification on the basis of that functional component. The majority of bio-stimulants in use today are complex mixtures of chemicals derived from a biological process or extraction of biological materials. The complexity of these mixtures is often considered to be essential to the performance of the bio-stimulant, and bio-stimulants may have properties of the whole, that cannot be fully elucidated by knowing the characteristics of the separate components or their combinations. This theory of complexity or “emergence” was described by Mayr ,planting gutter who argued that in many biological systems “the properties of the whole cannot be fully elucidated by knowing the characteristics of the separate components or their combinations.” “The term emergence describes the onset of novel properties that arise when a certain level of structural complexity is formed from components of lower complexity.

In the last few decades, emergence has been discussed in a number of different research fields, such as cybernetics, theory of complexity, artificial intelligence, non-linear dynamics, information theory, and social systems organization” . “Emergence” and “emergent properties” are thus closely related with the notion of the “systems biology” . Emergence was described by Johnson as “unexpected behaviors that stem from interaction between the components of an application and their environment,” “there is, however, considerable disagreement about the nature of ‘emergent properties.’ Others refer to emergent properties when an application exhibits behaviors that cannot be identified through functional decomposition. In other words, the system is more than the sum of its component parts” . Thus, a bio-stimulant could also be defined as “a formulated product of biological origin that improves plant productivity as a consequence of the emergent properties of its constituents.” To our knowledge, however, there have been no clear demonstrations that any bio-stimulant exhibits truly emergent properties. This is not however a unique challenge and all “biological systems are extremely complex and have emergent properties that cannot explained, or even predicted, by studying their individual parts” . Emergent properties have been demonstrated in the networks of biological signaling pathways ; in system-level study of traditional Chinese medicine , and in microbial communities . To adequately explain the biological complexity present in plants and their interactions with the environment, Lüttge and Bertolli et al. emphasize that classic reductionist biology/chemistry is indeed insufficient. While the two theoretical definitions provided in this section share a requirement that the mode of action is unknown, they differ in the core assumption that bio-stimulant function is a consequence of the discrete components in the bio-stimulant or as a consequence of the “emergent” properties of the bio-stimulant as a whole.

Each of these definitions is also incomplete in that it is certainly possible that a bio-stimulant may contain several molecules that act synergistically while not being truly “emergent,” and it is indeed possible and indeed likely, that even if a bio-stimulant is demonstrated to have emergent properties, that not all components of that bio-stimulant are required for that property to be expressed. We propose, therefore, a definition of a bio-stimulant that integrates these two concepts. Thus, a bio-stimulant is defined here as: “a formulated product of biological origin that improves plant productivity as a consequence of the novel, or emergent properties of the complex of constituents, and not as a sole consequence of the presence of known essential plant nutrients, plant growth regulators, or plant protective compounds.” Consistent with this definition, the ultimate identification of a novel molecule within a bio-stimulant that is found to be wholly responsible for the biological function of that bio-stimulant, would necessitate the classification of the bio-stimulant according to the discovered function. A review of the history of bio-stimulants and related products provides insight into the diversity of these products and the development of this field of study. The evolution of bio-stimulant classifications as described by various authors is presented in the Table 2. To the best of our knowledge, one of the first attempts to categorize bio-stimulants was provided by Filatov when 4 groupings of biogenic stimulants were suggested. Karnok compiled a list of 59 materials presenting in 15 bio-stimulants; Ikrina and Kolbin systematized patent literature and specified 9 categories of natural raw materials used to derive bio-stimulants; Basak suggested that bio-stimulants could be grouped on the basis of single or multi-component formulations and classified on the origin of the active ingredient, and the mode of action of the active ingredient. Du Jardin developed a scientific rationale of classification considering 8 categories of bio-stimulants and subsequently reduced this list to 7 categories .

Du Jardin was explicit in his exclusion of microorganisms from his categorization primarily to avoid conflict with existing categorization of microorganisms as bio-pesticides and sources of plant hormones. Later Bulgari et al. proposed a bio-stimulant classification on the basis of their mode of action rather than on their composition. Many bio-stimulant products have been classified into completely divergent groups and categories of function, use, and type of activity . For example, humate based products are often described as soil health amendments while plant growth promoting rhizobacteria could be categorized as bio-fertilizers, phytostimulators, and bio-pesticides . Du Jardin has proposed that bio-fertilisers are a subcategory of bio-stimulants. Seaweed extracts have been considered as bio-fertilizers and microorganisms have also been described as bio-fertilizers . Some inorganic elements or small molecules that are not known to be essential may also be classified as bio-stimulants if evidence of plant growth promotion is available . Thao and Yamakawa , for example, consider phosphites to be bio-stimulants since plant response to phosphites frequently cannot be explained as a consequence of the known anti-fungal function of these molecules. While the categorization of bio-stimulants by their origin does not, a priori, provide information on their mode of action this categorization may still be a useful tool to aid in the process of discovery and facilitate comparison between similar products. Registration of products used in agriculture is crucial to ensure their practical, safe and legitimate application. In the absence of a sound definition of bio-stimulants as a discrete group of products , the registration procedure and subsequent classification regime is untenable and this inevitably creates a barrier to trade and development. Various countries, states, and administrative regions have developed different categories for registration of potential bio-stimulants including terminology such as plant conditioners, “other fertilizers,” supplements, soil improvers, gutter berries plant strengtheners, fitofortificants, etc. . In many jurisdictions regulatory practices require an itemized description and identification of substances in all commercial product classifications while in others the registration of non-fully identified substances is allowed if those products are considered of complex composition. There is even a proposal for complex bio-stimulants to not specify the chemical name and note as “None” with the definition that “this product is a complex mixture of chemical substances” . If we accept the concept that a bio-stimulant is a product of clear benefit but unknown mode of action, then it can only be regulated by its safety and proof of efficacy. For example, in pharmacology it has been suggested that “the demand to demonstrate the mode of action of each single component in a phytopharmaceutical may not be obligatory any more” . The complex multi-component nature of many bio-stimulants clearly complicates discovery of their modes/mechanisms of action, production, registration and use. What is clearly needed however, is a regulatory mechanism to ensure that the products are “generally recognized as safe,” have “a positive benefit on crop productivity” and are discrete from exisiting categories of products. The task of identifying function and agronomic utility can then be pursued independently and will be driven by the marketplace imperative for product quality and consistency. Coordinating national legislation within this framework will become critical for the optimization of bio-stimulants and trade between different countries. The possible place of bio-stimulants in the regulatory system of pesticides and agrochemicals is illustrated in Figure 1.We have conducted an exhaustive analysis of the literature and categorized the majority of the reported bio-stimulants by origin . Microorganisms are widely used for the production of bio-stimulants and may be derived from bacteria, yeasts, and fungi. These preparations may include living and/or nonliving microorganisms and their metabolites. The concept of microorganism-based preparations as bio-stimulants is described by Xavier and Boyetchko , Sofo et al. , Colla et al. , Matyjaszczyk , and Ravensberg . different species of algae, mostly seaweeds, are also commonly used for producing bio-stimulants.

Seaweed-based preparations as bio-stimulants are described in reviews by Crouch and van Staden , Khan et al. , Craigie , Sharma et al. ; and experimental papers by Goatley and Schmidt , Jannin et al. , Billard et al. , Aremu et al. . Raw materials for bio-stimulants are also commonly based on higher plant parts including seeds, leaves, and roots and exudates from families Amaryllidaceae, Brassicacae, Ericaceae, Fabaceae, Fagaceae, Moringaceae, Plantaginaceae, Poaceae, Rosaceae, Solanaceae, Theaceae, Vitaceae, among others . bio-stimulants may also be based on protein hydrolysates and amino acids of animal origin including wastes and by-products , and insect derived chitin and chitosan derivatives . Humate-based raw materials are widely used to derive bio-stimulants and have been reviewed by Sanders et al. , Kelting et al. , Ertani et al. , and Jannin et al. . A final category of bio-stimulants includes those derived from extracts of food waste or industrial waste streams, composts and compost extracts, manures, vermicompost, aquaculture residues and waste streams, and sewage treatments among others. Because of the diversity of source materials and extraction technologies, the mode of action of these products is not easily determined. The technologies used in the production and preparation of bio-stimulants are highly diverse and include cultivation, extraction, fermentation, processing and purification, hydrolysis, and high-pressure cell rupture treatment . In some instances, a bio-stimulant product may also contain mixes of components derived from different sources and production methods. Frequently the rationale for utilizing extracts rather than raw biomass is a consequence of the need for a standardized manufacturing process to produce a uniform commercial product . For many products, the production processes are driven by process and marketing demands and are not the result of a targeted strategy to optimize the biological efficacy of the commercial product. While the ultimate composition and possible function of commercial bio-stimulant products may be partially determined by both the source of raw material and the process by which it is prepared , there may be manufacturing processes and product treatments utilized that result in compounds that are not present in the initial material. An example of this is the multitude of commercial seaweed extracts, often derived from the same species, that are rarely equivalent . Commercial bio-stimulant manufactured from similar sources are usually marketed as equivalent products, but may differ considerably in composition and thereby in efficiency . Many manufacturers do not reveal the technology of bio-stimulant production since that is a commercial secret .A diversity of substances contained in raw materials is used for the production of bio-stimulants. Whereas, primary metabolites are contained in most preparations de facto, the presence of secondary metabolites is more specific and depends to a large extent on the raw material used . Primary metabolites include amino acids, sugars, nucleotides, and lipids . Secondary metabolites are formed from different primary metabolic pathways, including glycolysis, the tricarboxylic acid cycle , aliphatic amino acids , the pentosephosphate and shikimate pathways which are primarily the source of aromatic AA and phenolic compounds , terpenoids/isoprenoids, nitrogen-containing compounds , sulfur-containing compounds. Frequently, bio-stimulants are shown to have a multicomponent composition and may include plant hormones or hormone-like substances, amino acids, betaines, peptides, proteins, sugars , aminopolysaccharides, lipids, vitamins, nucleotides or nucleosides, humic substances, beneficial elements, phenolic compounds, furostanol glycosides, sterols, etc. .