Anti-nutritional factors in plant-derived milk: a review on effects and effective reduction methods

1 Introduction

Milk, a fundamental component of human nutrition, is derived from various sources, including human, animal, and plant origins. Each type has unique compositional and functional traits. Human milk is uniquely suited to fulfill the developmental requirements of newborns, providing vital macronutrients and bioactive substances that promote immunity and growth (Antunes et al., 2022). Animal-derived milks, especially cow’s milk, are extensively consumed worldwide and are known for their comprehensive nutritional composition, which includes high-quality proteins and essential micronutrients (Karoui and Bouaicha, 2024). Plant-based milk alternatives (PBMAs) have become increasingly popular due to dietary preferences, lactose intolerance, and environmental factors; however, their nutritional composition varies considerably based on the source and fortification methods (Kovanen et al., 2025).

Cow’s milk is a nutrient-rich beverage, providing approximately 3.3 g of protein, 3.4 g of fat, and 4.8 g of carbohydrates per 100 mL. It serves as a substantial source of essential micronutrients, providing calcium (120 mg), phosphorus (93 mg), iodine (16 µg), riboflavin (0.18 mg), and vitamin B12 (0.4 µg) per 100 mL. The protein in cow’s milk is of high quality, containing all essential amino acids, with a digestible indispensable amino acid score (DIAAS) above 100% for all essential amino acids (Duraiswamy et al., 2023). Owing to its lactose content and fat composition, cow’s milk has a pleasantly sweet and creamy flavor profile that adds to its sensory appeal (Walther et al., 2022).

Plant-derived milks serve as alternatives to animal milk. These drinks are prepared by blending water with plant ingredients (Silva et al., 2022). These drinks are expected to resemble cow’s milk in flavor profile, shelf life, appearance, and texture rather than being identical to those of cow’s milk. The growing demand for plant-based alternatives stems from their lower environmental impact compared with animal-based food products. These products are useful because they are often lower in calories than cow’s milk and free from cholesterol, lactose, and milk proteins. Consequently, many consumers believe that these alternatives can help them achieve wellness and fulfill their health objectives (McClements et al., 2020). Because of their favorable fatty acid profiles and antioxidant activity, plant-based alternatives, when consumed as part of a balanced diet, may help reduce the risk of developing heart disease, cancer, diabetes, and atherosclerosis (Prytulska et al., 2021).

However, the presence of anti-nutritional factors (ANFs) in plant-based milks (PBMs) poses significant challenges to their nutritional efficacy. ANFs are naturally occurring substances in plants that hinder the absorption and utilization of vital nutrients (Reyes-Jurado et al., 2023). Plant-based milks commonly contain ANFs such as phytic acid, tannins, saponins, lectins, and protease inhibitors. These compounds can chelate minerals such as calcium, iron, and zinc, diminishing their bioavailability and potentially resulting in deficiencies, particularly in populations that predominantly consume plant-based diets (Samtiya et al., 2020). These ANFs, in turn, are classified as anti-protein and anti-mineral factors, and they can negatively affect human health through their toxic effects, unpalatability, inhibition of growth, and digestive disturbances (Bester, 2023). Plant-derived anti-nutrients have been noted to cause a reduction in the in vivo and in vitro bioavailability of zinc and iron through direct interactions with minerals and micronutrients (Manzoor et al., 2021a).

Several processing techniques have been investigated to reduce ANF levels in plant-based milks and address these difficulties. Conventional methods for reducing ANF levels, including soaking, fermentation, germination, and heat treatments, have demonstrated varying degrees of efficacy. For instance, germination stimulates endogenous enzymes that break down ANFs, whereas fermentation can degrade phytic acid and increase mineral bioavailability (Singh and Arora, 2023). Heat treatments, such as boiling and autoclaving, help denature protease inhibitors and lectins, making proteins more accessible for digestion (Vikram et al., 2020). These techniques enhance the nutritional profile of PBMAs either individually or in combination. Figure 1 shows the integrated nutritional and antinutritional profile of major plant-based food sources. Relative levels represent a qualitative synthesis of published literature and may vary with cultivar/genotype, growing conditions, processing conditions, and analytical methodology (Karoui and Bouaicha, 2024; Das et al., 2025; Semwal et al., 2025; Singh et al., 2025; Yılmaz Tuncel et al., 2025).

Even though research efforts are continuously emerging, the prevalence and nutritional significance of ANFs found in the beverage matrix are relatively understudied compared with those in PBMA raw materials, particularly with regard to processing-induced changes to ANFs, ANF bioavailability, and differences between PBMA raw materials, laboratory-prepared PBMAs, and commercially available PBMAs. Here, we review this understudied topic by connecting the occurrence and biological activity of common ANFs to their changes during processing, nutritional and technological trade-offs, and impacts on commercial PBMA development. Reduction of ANFs is discussed not only from the standpoint of deactivating toxic components but also in terms of how reduction may impact nutrient bioavailability, retention of other desirable molecules, processing efficiency, and overall sustainability. Finally, the discussion of ANFs is placed into the context of SDG 2 and SDG 12 to emphasize important connections between ANF occurrence/prevalence, processing-induced changes, nutritional impact, and technological performance during PBMA development (Silva and Smetana, 2022).

While previous reviews have covered anti-nutritional factors (ANFs) in edible plants and pulses as well as the nutritional and technological factors that influence PBMAs, here we offer a matrix-specific perspective that connects ANF presence to its processing behavior during PBMA manufacturing and its subsequent nutritional implications. Special consideration is given to data gathered from whole foods or raw plant materials compared with in vitro-prepared PBMAs and commercially available beverages, as concentrations and therefore the nutritional impact of ANFs can vary between matrices. Further, quantitative data demonstrating reduction of ANFs across traditional and novel processing platforms are discussed with regard to overall nutrient retention, bioavailability, process parameters, scalability, economic implications, sensory outcomes, and environmental impact. Factors that impact the nutritional importance of remaining ANFs such as mineral–phytate balance, ANF–ANF interactions, processing history, bioaccessibility, gastrointestinal modification, and microbiota are also taken into account. Overall, we hope to connect these factors and move beyond discussing ANFs or processing steps individually to better understand how ANFs are managed during PBMA production and application.

2 Literature search and study selection

A systematic literature search was carried out to identify relevant articles reporting the occurrence, nutritional relevance, processing-induced reduction, and bioavailability of ANFs in PBMAs. Search engines included ScienceDirect, Web of Science, PubMed, and Google Scholar databases. Keywords were used in combination to include general terms related to the product matrices, target compounds of interest, and processing approaches as follows: (“plant-based milk” OR “plant-based milk alternatives” OR “plant milk”), AND (“anti-nutritional factors” OR “antinutrients” OR “phytic acid” OR “phytate” OR “tannins” OR “saponins” OR “lectins” OR “oxalates” OR “trypsin inhibitors” OR “protease inhibitors” OR “goitrogens” OR “cyanogenic glycosides”) AND (“processing” OR “soaking” OR “germination” OR “blanching” OR “cooking” OR “microwave” OR “extrusion” OR “irradiation” OR “fermentation” OR “enzymatic treatment”) OR (“bioavailability” OR “bioaccessibility”), OR “commercial plant-based beverages”). Literature published between 2010 and 2026 was considered, with particular emphasis on the most recent articles published between 2020–2026. Peer-reviewed original articles reporting information on ANF occurrence, quantitative changes during processing, nutritional effects, mineral bioavailability/bioaccessibility, or methods and characteristics of laboratory-prepared or commercially available PBMA products were prioritized. Review articles were also considered to provide context and mechanistic background. Articles were removed from consideration if they did not pertain to plant-based milk or alternative plant-derived matrices, did not offer information related to ANFs or their nutritional impact, or were unrelated to processing, bioavailability, or nutritional impact. Evidence was then organized based on whether the material under investigation was considered a raw plant ingredient, laboratory-prepared PBMA, or commercially available PBMA, and was qualitatively summarized by ANF of interest, processing method, reduction achieved, if applicable, nutritional impact, and/or technological relevance.

3 Nutritional comparison across plant-based milk

PBMAs are increasing in popularity owing to dietary restrictions and environmental considerations. Nonetheless, their nutritional compositions vary markedly from that of conventional cow’s milk. Cow’s milk generally possesses elevated concentrations of energy, fat, carbohydrates, vitamins C, B2, B12, A, biotin, pantothenic acid, calcium, phosphorus, and iodine compared with the majority of plant-based beverages. Soy beverages, despite offering relatively high protein and elevated levels of vitamins B1, B6, folic acid, vitamins E, D2, K1, magnesium, manganese, iron, and copper, may still lack a comprehensive nutrient profile. The protein concentration in the majority of milk replacements is ≤ 1%, rendering them less dependable as protein sources. Cow’s milk has a higher Digestible Indispensable Amino Acid Score (DIAAS) than most plant-based beverages, reflecting its superior protein quality. Consequently, despite fortification efforts, plant-based beverages do not fully match cow’s milk (Jeske et al., 2017).

PBMAs have become viable alternatives for consumers who prefer dairy-free beverages, providing potential health advantages and nutritional sufficiency when appropriately formulated. Specific PBMAs, particularly those sourced from soy and legumes, naturally possess elevated protein concentrations and, when fortified, can deliver vital vitamins and minerals similar to those found in dairy milk (Chalupa-Krebzdak et al., 2018). Recent research has indicated that specific PBMAs, especially those sourced from legumes such as soy and peas, are abundant in critical nutrients and bioactive compounds. These encompass isoflavones, phytosterols, and dietary fiber, which have been linked to numerous health benefits, including enhanced lipid profiles and glycemic regulation (Paul et al., 2020). Furthermore, advances in food processing technologies have facilitated the fortification of PBMAs with vitamins and minerals, thereby improving their nutritional profiles so that they more effectively meet dietary needs (McClements et al., 2020).

Table 1 summarizes the nutrient composition of selected plant-based milks, which is important for understanding their dietary value. Almond milk is characterized by low calorie and fat contents, but also a low protein content. Soy milk has high amounts of protein and essential amino acids. Oat milk has higher carbohydrate and fiber contents, but moderate protein and fat levels. Coconut milk is high in saturated fats but low in protein, while rice milk is known to be hypoallergenic and higher in carbohydrates but low in protein. Hemp, tiger nut, and sesame seed milks represent newer options with unique nutrient profiles that may support various dietary requirements.

TABLE 1. Different plant-based milk nutrition and health benefits.

3.1 Commercial formulation and processing of plant-based milk alternatives

The content and nutritional significance of ANFs will also vary according to processing conditions and the formulation used to prepare the finished beverage. Raw plant ingredients, laboratory-prepared PBMAs, and commercially formulated beverages should therefore be considered separately (Sharma et al., 2025). While raw cereals, legumes, nuts, and seeds can contain relatively high concentrations of ANFs, these concentrations should not be expected to translate directly to the finished beverage, as soaking, grinding/aqueous extraction, filtration, thermal treatment, fermentation, and homogenization have all been shown to impact ANF concentration, chemical form, and bioaccessibility (Savitri et al., 2022). Importantly, evidence supporting individual ANFs is not uniformly derived from finished PBMAs. While studies have specifically examined plant-based milk/beverage matrices for some ANFs (e.g., phytic acid, tannins, saponins, lectins, and trypsin inhibitors), evidence supporting other ANFs has been extrapolated from raw plant ingredients or other related food matrices, rather than from finished PBMAs (Kuksal et al., 2026; Abbou et al., 2025). Evidence from these studies can inform our understanding of how ANFs may occur and function but cannot be assumed to demonstrate similar concentrations or nutritional impacts within the finished beverage product. Therefore, whenever possible, results obtained from raw plant ingredients, laboratory-made PBMA comparisons, and commercial PBMAs are differentiated throughout this review.

Compared with commercially produced beverages, PBMAs prepared in laboratories typically undergo fewer formulation and stabilization steps and are most often prepared using plant material and water alone (Reyes-Jurado et al., 2023). Commercial PBMA products may undergo several industrial operations to ensure microbial safety, physical stability, and desirable texture, shelf life, and sensory acceptability. Commercial products may also contain vitamins and minerals, added oils, emulsifiers, stabilizers, sweeteners, flavors, and other functional ingredients. While fortification may help improve nutritional adequacy, enrichment of PBMAs with minerals and vitamins will not necessarily negate the effects of residual ANFs (Cawthorne, 2025). For example, interactions between added minerals and residual phytate and other chelating ANFs could impact mineral bioavailability.

In summary, the presence of an ANF in a raw ingredient should not be assumed to carry the same nutritional significance as exposure from a commercial beverage. Conversely, measurement of a low ANF concentration in a laboratory-prepared beverage cannot always be extrapolated to commercially produced beverages with varying plant-to-water ratios, extraction efficiencies, processing conditions, fortification levels, and stabilizing ingredients. Direct chemical analysis of finished commercial PBMA products is therefore necessary to determine actual concentrations and the nutritional significance of residual ANFs. Special consideration should be given to this when interpreting mineral bioavailability and assessing nutritional quality across categories of PBMAs. ANF concentrations are expected to vary according to plant genotype, raw-material composition, plant-to-water ratio, extraction efficiency, processing conditions, formulation, and fortification. Direct comparisons should therefore be based on finished beverage measurements whenever available (Table 2).

TABLE 2. Distinction among raw plant materials, laboratory-prepared and commercial plant-based milk alternatives.

3.2 Anti-nutritional factors in plant-based milks

Plant-based diets have gained increasing popularity due to their environmental and health advantages. The body’s capacity to absorb and use nutrients may be hampered by substances called ANFs, which are present in many plant-based foods (Azra et al., 2023). ANFs also decrease the uptake of both macro- and micronutrients (Tufail et al., 2024). Antinutrients are mostly found in beans/legumes, nuts, and grains, although they can also be found in the leaves, roots, and fruits of other plant sources (Popova and Mihaylova, 2019). Common antinutrients found in plant sources include those depicted in Figure 2. The data presented in this summary vary in matrix specificity. Ideally, we use data collected directly from plant-based milk alternatives. Unfortunately, limited data exist for some ANFs and we relied on data collected from raw plant sources or other plant-based food matrices. These findings are interpreted as indicating the potential occurrence of ANFs rather than serving as direct measurements of ANFs in final PBMAs.

3.2.1 Saponins

Saponins are widely distributed throughout nature, with most reported sources being plants; they are non-volatile, surface-active compounds derived from secondary metabolism. Saponins have a sugar moiety in their structures and are classified as steroidal or triterpenoid compounds (Samtiya et al., 2020). Triterpenoid saponins have been isolated from several important legumes, such as beans, peas, and soybeans. Other plants in which triterpene glycosides have been identified include ginseng, tea leaves, spinach, quinoa, and sugar beet. Steroid saponins have been found in tomato seeds, Capsicum peppers, asparagus, and oats, among others (Zandona et al., 2021). Saponins are found in soy milk (soyasaponins), oat milk (avenacosides), and pea milk (Molina et al., 2025).

Harmful effects: Saponins remain in the gastrointestinal tract (GIT) for a prolonged period and are poorly absorbed; therefore, the processes that occur during digestion are closely linked to their bioactivities (del Hierro et al., 2018). The bioavailability of saponins is greatly influenced by their solubility in the gastrointestinal tract, as water makes up a significant portion of intestinal contents. Because of their amphiphilic nature and distinctive glycosidic groups, the majority of saponins are highly water-soluble and can self-micellate in aqueous conditions (Timilsena et al., 2023). In extreme cases, gastrointestinal damage, inflammation, or lesions may appear, which could allow saponins to enter the bloodstream. When saponins enter the bloodstream, they can cause hemolysis, seizures, liver problems, coma, and respiratory failure (Sharma et al., 2023).

Potential benefits: Foods rich in saponins may significantly contribute to lowering plasma cholesterol, preventing osteoporosis and ulcers, and diminishing the risk of cardiovascular disease. They may also reduce the incidence of coronary heart disease in males (Salim et al., 2023).

3.2.2 Lectins

Lectins, also referred to as hemagglutinins, are glycoproteins or proteins containing one or more non-catalytic domains that bind reversibly to specific mono- and oligosaccharides. Without altering the properties of the carbohydrates, they can adhere to the sugar molecules on erythrocyte surfaces and agglutinate them (Singh and Arora, 2023). Lectins are present in various plant-based milks, particularly those derived from legumes and cereals. They are found in soy milk, chickpea and lupin milks, and oat milk (Adamcova et al., 2021).

Harmful effects: Lectins can impair the GIT absorption of monosaccharide units, which are the products of the breakdown of carbohydrates. They can bind to the carbohydrate moieties that are exposed on the luminal cell membranes of the GIT, thereby interfering nonspecifically with the absorption of nutrients (Lyngdoh and Sharma, 2023). Additionally, lectins have systemic effects that include increased protein catabolism, disturbance of mineral metabolism, and breakdown of glycogen and stored fat (Veer et al., 2021), as summarized in Table 3.

TABLE 3. Types of antinutrients and their effect on nutrition.

Abbreviations used are: HCN, Hydrogen cyanide.

Health benefits: Lectins play various biological roles in both microorganisms and animals, including functions in innate immunity, regulation of the cell cycle, production of glycoproteins, and protection against infections (Lucius, 2020). Since lectins are known to affect cancer cells, they can be used to treat and identify the disease (Gupta et al., 2022).

3.2.3 Phytic acid or phytates

Many plant tissues, especially those of the Poaceae family (such as barley, rice, rye, and wheat) and legumes, predominantly accumulate phosphorus in the form of phytic acid, sometimes referred to as inositol hexaphosphate (IP6) or phytate. It is a significant antinutritional component (Ram et al., 2020). Phytates are found mainly in soy and oat milks (Daryani et al., 2024).

Harmful effect: Phytic acid is present in many plant-based foods, constituting 1%–5% of the weight of various cereals and legumes. Its presence in food raises concerns because research shows that it decreases the bioavailability of certain critical minerals through the formation of complexes with proteins and/or multivalent cations that may be insoluble or otherwise inaccessible under physiological conditions (Mihrete et al., 2019). Phytate has a strong affinity for chelating Zn²⁺, Fe²⁺, Mg²⁺, Ca²⁺, Mn²⁺, Cu²⁺, and K⁺ both in the human body and in food (Figure 3). Consequently, minerals chelated by phytic acid lack bioavailability, posing a public health issue because low mineral intake and limited food availability ultimately lead to compromised micronutrient status, increased mortality, and stunted growth (Brouns et al., 2022).

Health benefits: However, when consumed in small amounts, phytic acid can also have some beneficial effects. These include lowering blood glucose levels, reducing the absorption and digestion of starches, preventing dental cavities, preventing colon cancer, and increasing antioxidant capacity (Rahate et al., 2021).

3.2.4 Tannins

Tannins belong to the polyphenol category. They are bitter and astringent chemical substances with molecular weights ranging from 500 to more than 3,000. They interact with and bind to proteins and other substances, such as amino acids and alkaloids, and exhibit broad biological activity. They are also resistant to heat (Singh et al., 2023). Tannins can bind proteins because of the presence of at least 12 hydroxyl groups and five phenyl groups. The two primary categories of tannins are hydrolysable tannins and condensed tannins. Condensed tannins consist of flavonoids such as flavan-3-diol and flavan-3-ol, which lack a sugar unit. Hydrolysable tannins are made up of sugar, primarily glucose, bonded to gallic and ellagic acids. They are mostly found in almond milk and soy milk (Das et al., 2022).

Tannins are frequently categorized as antinutritional agents, because they can bind proteins and enzymes or obstruct the absorption of iron and minerals. Tannins may affect the body in two ways: low-molecular-weight tannins and microbial metabolites may have systemic impacts in different organs, whereas larger, complex structures that are not absorbed may function within the digestive system with antinutritional and antimicrobial qualities (Kawabata et al., 2019). In addition to proteins, tannins can bind components involved in the absorption of pectins, proteins, and carbohydrates; therefore, digestive enzymes may be unable to act on them, which can reduce the nutritional value of the food. When tannic acid is used in excess, it can cause liver damage, nausea, vomiting, and abdominal pain, as shown in Figure 4.

Health benefits: Their benefits mainly include immunomodulatory action, probiotic and prebiotic effects, and protection against infections. They may help protect against osteoporosis, coronary heart disease, and cancer. Tannins are utilized in industry as natural preservatives and are considered to confer health benefits (Tong et al., 2022).

3.2.5 Oxalates

A dicarboxylic acid called oxalic acid (C₂H₂O₄) is present in the cell sap of plants that are members of the Oxalis and Rumex species, either as potassium or calcium salts. The insoluble form of oxalic acid, calcium oxalate, is not readily absorbed once it passes into the digestive tract and is therefore excreted primarily in feces rather than urine (Tong et al., 2022). The highest oxalate content is found in almond milk (Borin et al., 2022).

Harmful effects: Oxalates are mostly present in plant foods such as grains, fruits, legumes, and vegetables. Oxalate is absorbed in the stomach, small intestine, and large intestine. Because insoluble oxalates have low bioavailability, they are eliminated in feces. Under normal circumstances, soluble oxalates constitute 5%–10% of the ingested oxalate and are absorbed by the colon and intestines (Borin et al., 2022). After being transported to the kidneys, free oxalate can either be excreted directly or combine with calcium ions to form calcium oxalate crystals, thereby raising urinary oxalate levels and potentially leading to significant health concerns, including kidney stones, a condition referred to as urolithiasis or nephrolithiasis (Bargagli et al., 2020).

Health benefits: Although oxalates are sometimes regarded as antinutrients because of their ability to bind minerals, certain oxalate-rich foods may provide protection against acidic chemicals that might cause dental erosion. This implies that oxalates may protect tooth enamel and promote dental health under certain conditions (Kumar et al., 2022).

3.2.6 Goitrogens

These are sulfur-containing glycosides, commonly associated with soybeans and Brassica vegetables. They are known to interfere with iodine uptake in the thyroid gland, potentially contributing to thyroid enlargement or goiter. The effects produced by goitrogens can be reversed by the iodine fortification of foodstuffs (Di and Giuliani, 2021). Plant foods such as Brassica vegetables are the main sources of dietary goitrogens known as glucosinolates, a large class of more than 120 compounds. Myrosinase, an enzyme present in plant tissues and produced by gut microorganisms, breaks down glucosinolates in damaged plant tissue after chewing and digestion, producing a range of compounds, including thiocyanates, nitriles, isothiocyanates, and sulforaphane (Kryczyk-Koziol et al., 2023). Goitrogens can be found in soy milk (Oruma et al., 2021).

Harmful effects: High concentrations of the thiocyanate ion can cause goiter, particularly when the diet contains little iodine. As a goitrogen, thiocyanate lowers thyroid function and causes goiter by blocking the thyroid gland’s iodide pump from absorbing iodine (Umar et al., 2021). Additionally, goitrogens can cause hypothyroidism. In hypothyroidism, goitrogens can increase thyroid enlargement and further suppress thyroid function. Similarly, goitrogens comprise a range of chemicals, medications, and foods that can interfere with thyroid function. This means that goitrogens are important not only for their role in inducing goiter but also for their potential to interfere with iodine metabolism overall. They also impact many regions of the body since they directly contribute to the synthesis and metabolism of thyroid hormones (Narale et al., 2024).

Potential health benefits: Some goitrogenic compounds and their breakdown products, such as isothiocyanates and sulforaphane, may have antioxidant and chemoprotective properties. However, excessive intake may impair thyroid function, particularly in individuals with low iodine status (Umar et al., 2021).

3.2.7 Trypsin inhibitors

Chemical inhibitors and protein inhibitors are the two basic categories into which inhibitors can be divided based on their characteristics. Protease inhibitors are proteins that inhibit protease activity. Trypsin inhibitors (TIs) are proteins that act as a kind of serine protease inhibitor by controlling the activation and interactions among proteins. This reduces proteolytic activity (Narale et al., 2024). Among plant-based milks, soy milk is the main source of trypsin inhibitors (Luo et al., 2025).

Harmful effects: The digestive enzyme trypsin is classified as a serine protease that is first synthesized as trypsinogen, an inactive precursor (Lale et al., 2023). At high levels, trypsin inhibitors form indigestible complexes with digestive enzymes and significantly restrict the activity of important pancreatic enzymes, including chymotrypsin and trypsin, thereby decreasing the digestion and assimilation of dietary proteins (Dey et al., 2020).

Potential benefits: Trypsin inhibitors have attracted interest for their possible health advantages. These inhibitors may exhibit anti-inflammatory and antibacterial characteristics, indicating their potential as bioactive molecules in medicinal applications. This viewpoint transforms the perception of trypsin inhibitors from mere dietary issues to possible contributors to health enhancement (Devi et al., 2019).

3.2.8 Cyanogenic glycosides

Cyanogenic glycosides are naturally occurring secondary metabolites found in plants. More than 2,500 plant species are known to possess cyanogenic glycosides. Flaxseed milk is a type of plant-based milk that can contain cyanogenic glycosides (Umar et al., 2021).

Harmful effects: When plant material containing these glycosides is ingested, β-glucosidase breaks them down, producing an aglycone and a sugar. Following this, hydroxynitrile lyase reacts with the aglycone to generate cyanide, an aldehyde, or a ketone. Death is among the most severe outcomes of cyanide’s toxicity. Cyanide prevents the body from synthesizing ATP by blocking cytochrome oxidase, the final stage of the electron transport chain, as shown in Figure 5. Rapid, deep breathing; an accelerated, arrhythmic, and weakened pulse; mouth frothing and salivation; muscle spasms; pupil dilation; and bright red mucous membranes are some of the symptoms that may occur before death (Narale et al., 2024). Cyanide released from cyanogenic glycosides accounts for their toxicity. Cyanide can be obtained through acid or base hydrolysis of cyanogenic glycosides. Despite its acidic environment, the stomach does not induce significant release of hydrogen cyanide (HCN) from cyanogenic glycosides (Vikram et al., 2020).

Potential benefits: High average concentrations of cyanogenic glycosides can be found in grain legumes, or pulses. However, the low amounts of cyanide usually present as intact glycosides, or cyanogens, are generally harmless in healthy, undamaged plant tissue because hydrolytic enzymes are not released unless tissue damage or decay occurs; when released, these enzymes can generate hydrogen cyanide (Luo et al., 2025).

3.2.9 Alkaloids

Alkaloids are one of the most extensive and structurally diverse groups of nitrogen-containing secondary metabolites produced by plants. They are characterized by a basic alkaline nature and typically include compounds such as quinine, caffeine, morphine, and nicotine (Lale et al., 2023). Certain alkaloids act as antinutrients. Solanine is an alkaloid that may cause gastrointestinal and neurological disturbances in individuals exposed to large amounts, approximately 20 mg/100 g, from potatoes. Several plant-based milk alternatives, such as those made from soy, almond, oat, and rice, have been shown to contain tropane alkaloids like atropine and scopolamine (Sinha and Khare, 2017).

Harmful effects: Although plant extracts may be consumed in very small quantities, plants containing harmful alkaloids can damage the human body. Through specific mechanisms involving enzymes, transporters, receptors, and genetic components in specific cells and tissues, toxic alkaloids might interfere with the normal functions of human physiological systems. The chronic effects include arrhythmias, psychosis, paralysis, sudden death, and teratogenicity. However, mild to moderate harmful symptoms of ingesting dangerous alkaloids include gastrointestinal upset, vomiting, itching, and nausea (Lamp et al., 2021).

Benefits: Alkaloids have been widely used in numerous traditional and modern medical applications, including cholinomimetic, anticancer, antiplasmodial, anti-asthma, antiarrhythmic, vasodilatory, analgesic, antibacterial/antibiotic, and antihyperglycemic pharmacological activities (Huang and Bu, 2022; Adibah and Azzreena, 2019; Abbas, 2020). An ideal balance between ANFs and nutrients helps to reduce negative impacts and improve health-promoting bioactivities. When taken in appropriate amounts, ANFs can provide health advantages since they show health-promoting actions, including the regulation of insulin responses, plasma cholesterol levels, triglyceride levels, and blood glucose, as shown in Figure 6 (Fraga-Corral et al., 2020; Molnar et al., 2024).

3.2.10 Matrix-dependent occurrence and relevance of ANFs in plant-based milk alternatives

The nutritional significance of ANFs is strongly shaped by the matrix. Plant ingredients may be naturally rich in phytate, tannins, saponins, lectins, oxalates, protease inhibitors, or other bioactive compounds; however, concentrations entering the consumer’s diet are influenced by all steps of the beverage-processing chain. Soaking can increase leaching and activate endogenous phytase, milling increases available surface area for extraction, filtration can remove insoluble ANFs, and heat treatment can denature proteinaceous ANFs such as lectins and trypsin inhibitors (Amin et al., 2022). Fermentation can further alter phytate and other ANFs through microbial enzyme production and metabolic activity (Kårlund et al., 2020). Thus, consideration of the nutritional impact of an ANF should be based on residual concentration and bioaccessibility in the finished beverage matrix instead of the absolute concentration in source ingredients.

This concept is especially important when considering commercial PBMs. Commercial products have generally undergone combinations of extraction, heat treatment, homogenization, stabilization, and fortification and may include ingredients that are not typically used to prepare PBMs in the research setting (Günan and Ömeroğlu, 2025). As such, two PBMs made from the same plant source may provide very different nutritional profiles and exposures to ANFs. Recent assessment of commercial PBMAs has revealed wide variability between product categories and formulations. Across 219 products from 21 brands, almond, oat, soy, coconut, rice, pea, hemp, flax, nut, and blended beverages were found to cover a wide nutritional range, with fortified soy products coming closest to matching the nutrient profile of dairy milk (Johnson et al., 2025).

Critically, commercial product labels typically do not report concentrations of ANFs such as phytate, tannins, lectins, or trypsin inhibitors. As such, a lack of ANF declaration on a product label should not be assumed to indicate the absence of these compounds. Analytical evaluation of finished commercial products is necessary to determine actual exposures. This highlights an important gap in the literature and underscores the need to develop and implement standardized analytical methods and subsequent reporting of ANFs in PBMAs.

For these reasons, we recommend that future studies report ANF concentrations on a common basis, e.g., per 100 mL or per serving, along with the raw-material concentration, processing conditions, dilution factor, fortification level, and, where feasible, in vitro gastrointestinal bioaccessibility. Such information would facilitate comparisons across laboratory and commercial preparations.

3.2.11 Factors modulating the real nutritional impact of ANFs: from presence to bioavailability

The presence of an ANF in a plant-based milk is not, by itself, an adequate indicator of its nutritional impact; the actual physiological consequence depends on the food matrix, co-occurring nutrients, and processing history. A more informative and increasingly standard approach is to express the antinutritional burden as a phytate-to-mineral molar ratio rather than as an absolute concentration. A phytate:zinc (Phy:Zn) molar ratio below 5 is associated with high estimated zinc bioavailability (∼30%), and a ratio above 15 with low bioavailability (∼15%); similarly, a phytate:iron (Phy:Fe) molar ratio below 1 (with 0.4 considered preferable) supports adequate iron absorption, while a phytate:calcium (Phy:Ca) ratio above 0.17 is associated with impaired calcium absorption. These thresholds provide a quantitative framework onto which the data can be mapped. For example, the study-specific reductions reported in Table 4 indicate that fermentation may achieve substantial phytate degradation in some soybean and maize-based systems, while soaking alone may have a more limited effect in certain pulse matrices (Samtiya et al., 2020).

TABLE 4. Selected literature examples of anti-nutritional factor reduction achieved by different processing treatments in plant-based milk matrices and related raw ingredients.

Reported reductions are study-specific and should not be interpreted as directly comparable across processing methods because plant source, matrix type, initial ANF concentration, processing conditions, raw-material-to-water ratio, and analytical methods differed among studies. Where initial concentrations were not reported in the cited study, baseline values are indicated qualitatively. Reductions reported for raw grains, legumes, seeds, or flours may not directly reflect reductions expected in finished laboratory-prepared or commercial plant-based milk alternatives.

Mineral fortification, a common strategy in commercial PBMs, does not automatically resolve this issue: because chelation is governed by molar ratios rather than absolute mineral content, adding calcium or iron to a beverage that retains high residual phytate can still leave the added minerals substantially unavailable for absorption unless the fortification level is calculated to offset the prevailing phytate:mineral ratio, or unless phytate is concurrently reduced through processing (Hurrell, 2004; Zhang et al., 2020).

Phytase activity is central to phytate reduction during soaking, germination, and fermentation. Ungerminated seeds contain low levels of endogenous phytase that are activated by hydration and mildly acidic, warm conditions; germination and pre-soaking substantially increase this activity, which is why germinated and pre-soaked substrates fermented with phytase-positive lactic acid bacteria, such as certain Lactobacillus and Bifidobacterium strains, achieve markedly greater phytate degradation than fermentation of unsoaked, ungerminated material. Strain-to-strain variability in phytase expression (as illustrated by the contrast between Bifidobacterium pseudocatenulatum- and B. longum subsp. Infantis-derived phytase in engineered soy-drink fermentations, achieving 90% versus 65% phytate hydrolysis, respectively) means that the choice of starter culture is itself a determinant of final bioavailability, not merely a matter of achieving acidification or shelf-life extension (Reale et al., 2007; Rollán et al., 2019).

ANFs also interact with one another rather than acting in isolation. Tannins can co-precipitate with phytate–protein complexes, and the combined presence of tannins and phytate can have a greater net inhibitory effect on mineral and protein bioavailability than either compound alone, while processing steps that reduce one ANF, such as dehulling, which removes the tannin-rich seed coat, frequently reduce others simultaneously because they share a common physical or enzymatic removal pathway. This means that single-ANF reduction data, while informative, can understate or overstate the net bioavailability benefit of a given treatment unless multiple co-occurring ANFs are considered together, as attempted in Table 5.

TABLE 5. Selected antinutritional factors: biosynthesis pathways, adverse effects, potential bioactivities, and reported intake guidance.

At the molecular level, protein–phenolic, i.e., tannin–protein, and mineral–phytate complexation are both strongly pH-, temperature-, and matrix-dependent. Tannin–protein binding is generally reversible and weakens under acidic gastric conditions and in the presence of proline-rich salivary or dietary proteins, whereas phytate–mineral complexes are least soluble near neutral pH and can re-form after cooking if the food is not further processed, for example, by fermentation, to hydrolyze the phytate backbone itself rather than merely disrupting the complex transiently (Cheryan and Rackis, 1980; Samtiya et al., 2020). Furthermore, because phytate-mineral binding is mediated by ion exchange, the presence of divalent cations in the food matrix can stabilize these aggregates, necessitating the use of specialized chelating agents or exogenous phytases to achieve meaningful reductions in mineral sequestration (Afify et al., 2011; Garrido-Galand et al., 2021).

Finally, gastrointestinal digestion and gut microbiota composition influence the terminal fate of residual ANFs after processing. Microbial phytase activity in the distal gut can hydrolyze a fraction of phytate that survives food processing and upper-gut digestion, partially recovering mineral bioavailability in a manner that varies with individual microbiome composition; similarly, colonic fermentation of unabsorbed tannin–protein complexes and saponins by gut bacteria can alter their systemic and local physiological effects, including some of the beneficial bioactivities, such as cholesterol-lowering effects of saponins and antioxidant effects of phenolic metabolites). Because these downstream, host- and microbiome-dependent processes are not captured by in vitro ANF assays, the values reported in Table 5 should be interpreted as indicators of processing efficacy rather than as direct proxies for in vivo bioavailability; the two are correlated but not equivalent, reinforcing why molar-ratio-based estimates and, where feasible, in vivo or Caco-2-based bioaccessibility assays provide a more complete picture of the nutritional relevance of ANFs in plant-based milks.

4 Methods for decreasing antinutritional factors

Plant components often contain antinutrients such as polyphenolic compounds, saponins, tannins, protease inhibitors, and phytate. These compounds can reduce protein digestibility and mineral absorption, cause health problems, and induce toxicity when present in high quantities, thereby interfering with the nutritional content of foods (Samtiya et al., 2020). Reducing antinutrients increases the absorption of specific cations such as Ca, Fe, and Zn and improves protein assimilation, which enhances the nutritional value of food (Mezdoud et al., 2022). Processing is one of the most important approaches for lowering the antinutrient content in food. Processing methods such as germination, boiling, dehulling, fermentation, soaking, and pressure cooking can reduce levels of antinutrients such as lectins, phytic acid, tannins, protease inhibitors, saponins, and phenolics (Table 5).

4.1 Dehulling

Dehulling is the process of removing the seed coat from pulses, which is an important post-harvest operation that enhances the palatability of food grains. However, it can also remove minerals and dietary fiber. Dehulling also removes the germ and the mucilaginous layer found between the hull and cotyledons. In other words, dehulling increases protein concentration while decreasing the levels of condensed tannins and polyphenols (Oruma et al., 2021). The typical components of pulse seeds include a seed coat and cotyledons. When soaked pulses are dehulled, the seed coat is typically removed and discarded. Discarding the seed coat removes some minerals as well as antinutrients (Thakur et al., 2019). Dehulling does not substantially reduce α-amylase inhibitors. However, dehulling pulses also seems to lower their tannin and polyphenol content (Borin et al., 2022).

4.2 Soaking

Soaking is an operation that involves immersing seeds or grains in water to reduce the time required for cooking. Soaking allows antinutrients to leach out of the seeds and into the soaking medium. Soaking thereby enhances digestion and makes nutrient absorption easier for the body (Hendek and Bektas, 2018). Soluble compounds such as IP6, tannins, TIs, raffinose-family oligosaccharides, and saponins migrate from the grains into the soaking water, thereby reducing associated negative health effects (Samtiya et al., 2020). The soaking technique involves completely immersing grains in water for a specified period to activate their natural phytases. Grains naturally contain endogenous phytases. These enzymes can be activated by a variety of treatments, such as soaking and other pretreatments, greatly enhancing the reduction of phytic acid levels. Soaking is among the common methods often practiced during the germination and fermentation of cereals. It is also used as a pretreatment for cereals before cooking (Rousseau et al., 2020).

4.3 Blanching

Blanching is the process in which food is dipped in boiling water with or without salt and allowed to boil for a few moments. It leaches out many antinutrients into the blanching water and inactivates several enzymes, including trypsin inhibitors, polyphenol oxidase, and others. The leaching of inhibitors into hot water may reduce trypsin inhibitor concentration (Samtiya et al., 2020; Purohit et al., 2023). Leaching mostly controls tannins, oxalates, and phytates in a standard blanching process. Higher processing temperatures can affect component diffusion in a favorable way. Blanching itself can disrupt cellular structures and enhance the leaching of antinutritional compounds. However, it is not as effective as a physical approach such as particle size reduction (Bhasin et al., 2018).

4.4 Germination

Germination is a traditional form of processing that is crucial for reducing levels of antinutrients, especially phytate, in grains and pulses (Patterson et al., 2017). Germination is characterized by high enzymatic activity that contributes to improved nutritional value and amino acid bioavailability (Abbas and Ahmad, 2018). Various studies have indicated that the process of germination is more efficient than heat processing for the elimination of phytic acid from pulses such as fava beans, peas, and chickpeas. Germination may also affect the levels of other antinutritional and specific bioactive components. Germination leads to the reduction of enzyme inhibitors, but not to the same extent as thermal treatment (Zhou et al., 2024). Moreover, some secondary metabolites, which act as reserve antinutrients, are mobilized during germination, largely owing to the reduction of ANFs (Dewan et al., 2024). Germination has one advantage over other methods, despite its moderate efficiency: it enhances certain characteristics and increases the concentration and bioavailability of various nutrients. If there is no time limit, germination could be combined with other methods that are at least as successful, including gamma irradiation. Combining the two strategies, for example, may help offset the vitamin C losses caused by gamma irradiation (Dey et al., 2020).

A comprehensive evaluation of PBMAs by Hu et al. (2022) highlighted the role of germination in lowering antinutritional components. The study found that prolonging germination time significantly reduces ANFs such as phytic acid and tannins. The activation of endogenous enzymes during germination degrades these substances. Germination improves mineral bioavailability and reduces nutrient-binding molecules in plant-based milks. Germination appears to be a promising, natural way to increase the quality and health benefits of PBMAs.

4.5 Cooking

Cooking reduces many antinutrients in vegetables, legumes, and whole grains. These include phytic acid, oxalic acid, and tannins. Since protease inhibitors are proteins, they are easily degraded by thermal processing. When lentils are cooked, the soluble solids contained in them are lost, and they become more concentrated in terms of protein and starch fractions (Devi et al., 2019). The application of hydrostatic pressure in high-pressure cooking serves to sterilize the food, denature proteins, and regulate enzymes and other biochemical reactions without causing physical deterioration of the raw material (Sharma, 2021). Cooking techniques are considered very effective for the denaturation of trypsin, chymotrypsin, and some other heat-sensitive molecules, thereby inactivating protein-based antinutritional agents. In Phaseolus vulgaris, this approach also eliminated 30%–40% of polyphenols (Abbas and Ahmad, 2018).

4.6 Microwave heating

Microwave heating is an advanced technology compared with conventional heating methods. It leads to rapid and uniform heating and can be very effectively applied to nonconductive food products. Microwave heating modifies the morphological, rheological, and functional properties of pulse starches, thus enhancing cooking quality and improving the dietary composition of pulses by reducing ANF levels (Chakraborty and Paul, 2019). As a dielectric thermal processing technique, microwave processing accelerates the inactivation of certain antinutrients compared with traditional processing methods. Losses of minerals and B vitamins in microwave-cooked chickpeas were lower than losses occurring during boiling and autoclaving (Devi et al., 2019). Microwave treatment has been shown to affect ANFs in foods such as buckwheat, horse chestnut, black soybean, velvet bean, and kabuli chickpea (Suhag et al., 2021; Saini and Morya, 2021).

4.7 Fermentation

Fermentation is an established method for improving the nutritional value of grains through the enhancement of grain digestibility. Carbohydrates are also oxidized during fermentation to generate energy. Fermentation can increase the levels of essential amino acids such as methionine, tryptophan, and lysine, which have been shown to improve grain quality (Mohapatra et al., 2019). The fermentation process may also improve the digestion and bioavailability of proteins, while microbial activity may lead to beneficial changes before ingestion (Ozolina et al., 2023). One initial step in this process may be the enzymatic breakdown of starch. In particular, lactic acid bacteria (LAB) are important during fermentation because they produce metabolites and enzymes, exhibit probiotic characteristics, contribute to the breakdown of cyanogenic glucosides, and support the formation of other bioactive molecules (Diouf et al., 2019). Phytate levels may decrease during fermentation due to microbial activity. During this time, reduced polyphenol activity may be caused by the presence of phenolic oxidases in bacteria (Arbab et al., 2021).

Kumari et al. (2022) studied the impact of lactic acid fermentation on the antinutritional components in soymilk. The researchers utilized Lactobacillus plantarum as a starter culture and monitored variations in phytate levels after fermentation. The research indicated that fermentation resulted in a substantial decrease in phytic acid concentrations, primarily due to the enzymatic action of phytase produced by the bacteria. This technique enhanced mineral bioavailability in soymilk, indicating that lactic acid fermentation is an effective method for improving the nutritional quality of plant-based beverages.

4.8 Dehydration

Dehydration leads to decreased antinutrient content in pulses. It causes an overall reduction in phytates, lectins, and enzyme inhibitors in pink-mottled beans, white beans, and lentils. Dehydration may enhance protein digestibility, rendering dehydrated pulse flours suitable as raw materials for the preparation of ready-to-use specialized meals (Ravichandran and Upadhyay, 2022).

4.9 Extrusion cooking

Extrusion is a thermal cooking technique increasingly utilized in food processing because of its efficiency, continuous operation, and high throughput (Kamau et al., 2020). The shear force used in the extrusion procedure deforms the complex structure, thereby denaturing trypsin inhibitors during extrusion cooking. Extrusion heating reduces antinutritional elements and increases food stability by deactivating the enzymes lipase and lipoxygenase (Bhullar et al., 2021). The effectiveness of the technique depends largely on the plant species, its genetic characteristics, and the temperature used (Faizal et al., 2023). In addition to increasing the digestibility of proteins and carbohydrates, extrusion significantly reduces antinutritional components such as phytic acid, lectins, trypsin inhibitors, and tannins (Zhou et al., 2024). Cooking legumes by extrusion has multiple benefits: it gelatinizes starch, increases soluble dietary fiber, reduces lipid oxidation, and helps maintain foods’ natural colors and aromas (Ciudad-Mulero et al., 2022).

4.10 Irradiation

Irradiation refers to the exposure of food to ionizing radiation, including alpha and beta particles, energetic electrons, gamma rays, or X-rays, for a specified duration (Lima et al., 2019). Irradiation can completely inactivate the lipoxygenase and trypsin inhibitor enzymes in soybeans (Joshi et al., 2020). However, phytic acid levels were slightly decreased after exposure to gamma irradiation, and the concentrations of tannins and phenols showed notable decreases. Gamma irradiation chemically breaks down phytic acid and glucosinolates. Maximum phytic acid removal was observed at doses exceeding 30 kGy (Kadam et al., 2021).

4.11 Roasting

Roasting is an important method for processing seeds. Overall, roasting greatly improves the appearance, texture, flavor, and color of seeds, thereby enhancing palatability. Moreover, roasting also eliminates undesirable microbes and inactivates enzymes, allowing the product to remain microbiologically safer (Zaaboul et al., 2019). This method may also help retain nutrients. It is a dry process, in contrast to wet cooking, which can cause nutrient leaching. Roasting reduces oligosaccharide content (Yegrem et al., 2021). Among the several benefits of the roasting technique are improved product quality, substantial roasting capacity, prolonged product shelf life, and reduced environmental impact (Bagheri et al., 2019). It also reduces the trypsin inhibitor activity in soybean meal (Shi et al., 2018). Roasting has been observed to eliminate many antinutritional or toxic components, including trypsin inhibitors, hemagglutinin, goitrogenic agents, cyanogenic glycosides, alkaloids, and saponins (Singh et al., 2018).

4.12 Enzymatic treatment

Enzymatic treatment is an efficient approach to reducing antinutritional factors (ANFs) in plant-based milks, thereby improving their nutritional quality. Enzymes such as phytase, protease, and α-galactosidase target specific ANFs, breaking them down into less harmful compounds (Smith et al., 2022). To lower ANFs in plant-based foods, Faizal et al. (2023) evaluated several food processing techniques, including enzymatic treatments (Joshi et al., 2020). The study demonstrated that enzymatic treatments efficiently reduce phytic acid, tannin, and enzyme inhibitor levels without degrading the food’s nutritional value. This method is particularly suitable for plant-based milks, where it is important to preserve both nutritional and sensory qualities.

4.13 Comparative evaluation of ANF-reduction technologies

Sections 4.1–4.12 describe individual processing methods, but a direct, criterion-based comparison is needed to guide method selection for plant-based milk (PBM) production (Table 6). No single technique is universally superior; rather, each involves trade-offs among effectiveness, nutritional quality retention, processing time, scalability, cost, and environmental footprint.

TABLE 6. Comparative evaluation of ANF-reduction processing technologies.

Wet, low-energy methods such as soaking are inexpensive and universally accessible but are largely ineffective against phytic acid and only partially effective against lectins and soluble oxalates, since phytic acid requires enzymatic hydrolysis, e.g., by phytase, or thermal cleavage rather than simple leaching. Germination and fermentation, by contrast, rely on endogenous or microbial enzyme activity and achieve some of the largest reported reductions in phytate, approximately 90%–97% with phytase-active lactic acid bacteria) and trypsin inhibitors, up to approximately 99%, while simultaneously enhancing free amino acids, B vitamins, and antioxidant capacity. Their principal drawback is time, ranging from hours to several days, and the need for microbiological or germination-condition control at scale, which raises process variability compared with thermal methods (Gupta et al., 2013; Kaushik and Singh, 2026).

Thermal processing, including boiling and autoclaving, remains the most universally applied industrial method and is highly effective against heat-labile protein-based ANFs, including lectins and trypsin/protease inhibitors, typically achieving greater than 90% inactivation. However, phytic acid, being thermally stable, is frequently unaffected or only marginally reduced by boiling alone. Extrusion cooking combines thermal and mechanical, i.e., shear, energy in a continuous, high-throughput format well suited to industrial PBM ingredient production; it achieves some of the highest reported reductions across multiple ANF classes simultaneously, including trypsin inhibitors up to 96%, tannins up to 92%, phytic acid up to approximately 57%. However, the elevated temperatures and shear can reduce lysine availability and cause losses of heat-labile vitamins, and the capital cost of extrusion equipment is higher than for soaking, germination, or fermentation (Avelar et al., 2021; Patterson et al., 2017).

Microwave treatment offers a middle path: it achieves ANF reductions comparable to or exceeding conventional wet heating such as 53%–65% reductions in phytic acid, polyphenols, and oxalates, and up to 100% reduction in saponins within 2.5 min, while generally preserving more heat-labile nutrients than boiling or autoclaving, because of shorter exposure times. Gamma irradiation can achieve near-complete phytate degradation at high doses (15–30 kGy) and substantial trypsin/chymotrypsin inhibitor reduction, with minimal impact on proximate composition at approved doses. However, it requires specialized, capital-intensive irradiation facilities and regulatory clearance, which restricts its practical accessibility for small- and medium-scale PBM producers and raises consumer-acceptance considerations in some markets (Das et al., 2025; Manikpuri et al., 2024; Sushil and Morya, 2026).

Enzymatic treatment, including the addition of targeted phytase, protease, or α-galactosidase addition, is the most ANF-specific approach and, because it acts on a single molecular target under mild conditions, generally causes the least collateral damage to desirable nutrients, flavor, and functional properties. Its main limitations are the added formulation cost of exogenous enzymes and the need for precise control of pH, temperature, and reaction time to achieve consistent, food-grade results at scale (Asghar et al., 2024; Woiciechowski et al., 2022).

Overall, the evidence indicates that combined (“hurdle”) approaches, for example, soaking followed by germination, or blanching followed by fermentation tend to outperform any single treatment, because each step targets a different resistance mechanism, namely, solubility-driven leaching, enzymatic hydrolysis, and thermal denaturation, respectively. For PBM manufacturers, the appropriate choice therefore depends on the dominant ANF in the raw material, the acceptable processing time and cost, and whether nutrient enhancement, as with germination/fermentation, or rapid, high-throughput processing, as with extrusion/microwave treatment, is the priority.

The quantitative reductions reported for each processing technology are specific to the studies from which they were obtained and are not intended to serve as directly comparable metrics of processing efficacy. Percentages were generated from different plant sources using a variety of raw-material-to-water ratios, initial ANF concentrations, processing temperatures and times, analytical methods, and study conditions. Therefore, the largest percentage reduction reported for an ANF using a particular processing technology is the maximal reduction observed under the conditions specified in that study, and may not be reflective of the reduction that can be expected during plant-based milk production. Furthermore, reductions seen in whole legumes, cereal grains, nuts, or seeds cannot be assumed to parallel reductions in laboratory-prepared or commercially available PBMAs, as extraction efficiency, filtration, dilution, formulation, fortification, and any additional processing can have significant impacts on the final concentrations of ANFs.

5 Future perspectives

Future PBMA processing strategies should aim to balance ANF reduction with the preservation of the nutritional and sensory attributes of PBMAs. For example, instead of simply minimizing ANFs, innovations should consider how to selectively minimize ANFs while maintaining or enhancing nutrients, bioactive components, sensory attributes, and processing efficiencies. Selective enzymatic treatments, such as phytase, protease, tannase, or α-galactosidase, may enable targeted hydrolysis of certain ANFs and could conceivably result in less nutrient loss than more intensive thermal processing methods. However, the efficacy of enzymatic treatments will depend on factors such as the specificity of each enzyme, other components of the processing regimen, the composition of the plant matrix, and suitability for industrial beverage production. Matrix-specific factors are also likely to affect enzymatic treatment effectiveness.

Another strategy may involve identifying plant varieties with inherently lower levels of ANFs. Through conventional breeding or marker-assisted selection, varieties with lower levels of phytate, tannins, lectins, or protease inhibitors could be explored as starting materials for PBMA production and may allow for less intensive downstream processing. Genome-editing techniques such as CRISPR may also facilitate genetic manipulation of metabolic pathways responsible for ANF biosynthesis in plants. However, it will be important for future research to consider how alterations in ANF levels may impact factors such as crop yield, nutritional quality, food safety, regulatory considerations, and consumer acceptance of products.

Moving forward, omics-based analyses could aid in discovering the metabolic pathways and mechanisms responsible for ANF biosynthesis and changes during food processing. Genomics, transcriptomics, proteomics, and metabolomics data could be leveraged to better understand which genes, proteins, and metabolites are associated with the accumulation of ANFs in plant matrices. Such data could also help identify genes and enzymes responsible for changes in ANF concentration during processing, and could aid in selecting ideal raw materials and tailoring processing conditions to each plant matrix.

Fermentation with selected microorganisms or microbial enzymes may represent another area of innovation for next-generation PBMA development. As fermentation can decrease certain ANFs, it may be possible to apply fermentation or controlled microbial bioprocessing to target the transformation of ANFs in PBMAs. In one study, fermentation of pearl millet with Lactobacillus reuteri decreased ANFs while increasing antioxidant capacity (Srivastava et al., 2024). However, scale-up, process control, production costs, regulatory barriers, and consumer acceptance of fermented PBMA products are important factors to consider with this approach.

In addition to balancing ANF reduction with nutritional and sensory quality, efforts to reduce ANFs should also take into account energy input and process sustainability. Many traditional food processing practices have been shown to reduce levels of ANFs in foods. For example, soaking, boiling, and roasting have been shown to reduce phytate content in some legumes and seeds (Godrich et al., 2023). Although these practices require additional time and resources, emerging processing technologies such as high-pressure processing, pulsed electric fields, ultrasound, and cold plasma show promise as non-thermal or mild-processing methods that could reduce ANFs while limiting energy input and processing time. Cold plasma, in particular, has been used to investigate its effects on the physical, nutritional, and sensory characteristics of milk and dairy products (Nikmaram and Keener, 2022).

Future work comparing the effects of different processing technologies on ANFs should also consider energy usage, water usage, processing time, generation of waste byproducts, nutrient loss, scalability, cost, and other factors related to sustainability and industrial applicability. Lastly, more efforts should be made to quantify residual ANFs in commercial PBMA products. Rather than relying on literature values reporting ANF concentrations in plant foods, direct analysis of finished PBMA products will allow for a better understanding of how different processing methods affect ANFs. In addition to measuring ANF concentrations, the impact of processing on mineral bioaccessibility, protein digestibility, gastrointestinal transformation, and overall nutritional quality should be considered. Taken together, these considerations can facilitate innovation in PBMA development, in which ANFs are managed alongside other considerations central to designing nutritious and sustainable plant-based foods.

6 Conclusion

Plant-derived milk alternatives offer significant nutritional benefits but face challenges because of ANFs that interfere with digestion and metabolism. Traditional methods such as soaking, fermentation, and thermal processing help mitigate ANF content, but emerging techniques such as targeted enzymatic treatment and advanced bioprocessing offer promising solutions. As consumer demand for sustainable alternatives grows, food scientists must refine ANF reduction strategies without compromising essential nutrients. Future research should focus on optimizing processing technologies, exploring synergistic detoxification approaches, and assessing the long-term health effects of residual ANFs to ensure that plant-based milk remains a viable and nutritionally enriched alternative to dairy products.

Generative AI statement

The author(s) declared that generative AI was not used in the creation of this manuscript.

Any alternative text (alt text) provided alongside figures in this article has been generated by Frontiers with the support of artificial intelligence and reasonable efforts have been made to ensure accuracy, including review by the authors wherever possible. If you identify any issues, please contact us.

Publisher’s note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

Footnotes

Edited by: Ana Belen Martin Diana, Instituto Tecnológico Agrario de Castilla y León, Spain

Reviewed by: Ni Ketut Sutiari, Udayana University, Indonesia

Josiane Domingues, Fluminense Federal University, Brazil