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Probiotic Lactobacillus casei Shirota and its Aflatoxin-Binding Properties
Probiotic Lactobacillus casei Shirota (LcS) has been extensively reported to have aflatoxin-binding ability. Here, we discuss the available evidence and mechanism of action for aflatoxin-reducing properties. The possible factors affecting the probiotic efficacy in removing aflatoxin are highlighted.

by Chang Wei Lin and Dr Mohd Redzwan Sabran


Probiotics are live microorganisms that have health benefits beyond basic nutrition (when consumed in adequate amounts). Lacticaseibacillus casei, more commonly known as Lactobacillus casei, is one of the most consumed and best characterised probiotics.

The name Lactobacillus casei has been officially reclassified into Lacticaseibacillus casei in April 2020, where its new name was derived from the combination of Latin words: “lacti” means milk, “casei” means cheese, and “bacillus” means small rod.1 It implies that the probiotic is a small rod-shaped microorganism that is often found in fermented dairy products.

The research into L. casei strain Shirota (LcS) began in the 1930s when the strain was isolated and cultivated by a Japanese scientist, Dr. Minoru. The Shirota strain was later developed into a new fermented milk drink named Yakult after five years of discovery.2

Aflatoxins and Its Toxic Effects

Aflatoxins are a family of toxins produced by certain moulds, particularly Aspergillus species. These moulds are abundant in warm and humid areas. There are four major aflatoxins, including aflatoxin B1 (AFB1 ), aflatoxin B2 (AFB2 ), aflatoxin G1 (AFG1 ), and aflatoxin G2 (AFG2 ). These aflatoxins are commonly found in food and feed such as nuts, spices, grains, and dried fruits. Herbal and traditional medicine may also be affected by aflatoxin contamination.3 Besides the B and G aflatoxins, aflatoxin M1 (AFM1 ) is the hydroxylated metabolite of AFB1 and is mainly excreted in milk or urine. When lactating animals ingested feed containing AFB1, these toxins are metabolised and excreted as AFM1 in milk. The contamination of AFM1 is not limited to raw milk but the whole milk chain since it could be carried over to dairy products.4 Dairy products such as cheese, cultured milk, and yoghurt have been detected with aflatoxins.5 These toxins cannot be seen with the naked eyes since they are colourless, odourless, and tasteless.

Acute exposure to a large dose of aflatoxins can lead to toxicity. This acute effect is characterised by abdominal pain, nausea, vomiting, and other signs of acute liver injury.6 These effects may worsen with high carbohydrate intake and low protein intake.7 Long-term consumption of aflatoxin-contaminated foods is a risk factor for liver cancer.

Aflatoxins have been classified by the International Agency for Research on Cancer (IARC) as carcinogenic to humans (Group 1), the same category as smoking and eating processed meat.8 Agents in Group 1 are known to have strong and clear evidence of carcinogenicity in humans. AFB1 is the most potent among all the aflatoxins. The relative potency of aflatoxins is reported in the order of AFB1 > (AFG1 and AFM1) >> (AFG2 and AFB2).9 Apart from being carcinogenic, AFB1 is known to be mutagenic, genotoxic, and immunosuppressive. Liew et al. (2022) proposed that the adverse effects are induced by aflatoxins by impairing the gut microbiota stability and increasing inflammation.10

Probiotic LcS as Potential Aflatoxin Binder

The aflatoxin-reducing properties of probiotics including LcS have been extensively discussed.11 Most animal studies were in favour of LcS to reduce aflatoxin levels and/or modulate the adverse effects of aflatoxins.12,13,14,15 These protective effects exist either by the administration of LcS individually or jointly with other components including chlorophyllin16 and a high protein diet.15 Indeed, AFB1-lysine adduct level in the blood samples of rats was reduced even when LcS was given prior to the aflatoxin exposure.17 This implies that the effects of treating LcS on pre- or post-aflatoxins exposure may be similar. It is plausible since LcS colonises in the gut for at least 14 days in adults18 or up to six months in children.19 Pre-exposure to probiotics may reduce the binding of aflatoxins with intestinal mucus, leading to faster removal.20 Thus, regular supplementation of probiotics may be of value for early prevention of aflatoxin toxicity, especially among those with a high risk of exposure.

A human interventional study exploring the effect of LcS in reducing aflatoxins is scarce. The first and only study was reported in a randomised, double-blind, cross-over, placebo-controlled trial, whereby 71 healthy adults were given fermented milk containing LcS followed by placebo drinks or vice versa for four weeks separated by a two-week washout period.21 The supplementation of LcS did not yield any conclusive findings on the reduction of serum AFB1-lysine adduct and urinary AFM1 due to several confounding factors. Nonetheless, findings from the literature based on in vitro and animal studies12,14,22 showed promising effects of LcS as aflatoxin binder.

Mechanism of Action

The aflatoxin-reducing properties of LcS are possibly attributed to its binding ability toward aflatoxin in the gut, which reduces the aflatoxin bioavailability.12 Aflatoxins adhere physically to the carbohydrate components of the probiotic cell wall by weak, non-covalent interactions corresponding to the formation of van der Waals interaction, hydrogen bonds, and electrostatic interactions.23 The aflatoxin-binding capacity of LcS is not limited to live cell but also other cell components although live cell was shown as the most efficient binder (98.0 per cent).12 The probiotics bound to aflatoxin are less likely to adhere to the intestinal wall, increasing the excretion of aflatoxin from the body.20 In addition to the aflatoxin-binding capacity, LcS may modulate the AFB1-induced gut microbiota imbalance and thus minimise the toxic effects of AFB1.22

The binding of probiotics towards aflatoxin is strain specific. AFB1-LcS complex was significantly more stable than other strains, retaining 93.8 per cent of AFB1 after four hours of incubation.24 The basis for the observed differences is unclear but it is speculated that the binding of aflatoxin may relate to the hydrophobicity of the cell surface.25 Gram-positive bacteria like LcS significantly removed more AFB1 than Gram-negative bacteria, indicating that the detoxicating effect depends on the structure of the cell wall.26 Under atomic force microscopy and scanning electron microscopy, aflatoxin binding has induced structural changes on the probiotic bacterial cell surface.12,17 The alteration in morphology may provide some hints that the binding of AFB1 occurs on the cell wall surface of probiotics. In addition, experimental data discovered teichoic acids as a key component of LcS cell wall structure that may be involved in complex binding.24 Further studies are needed to further elucidate these mechanisms.

Factors Affecting the Efficacy of Probiotic LcS

The binding activity of LcS towards aflatoxins can be affected by the gut condition, including gastric pH, digestive enzymes, and intestinal mucus. Probiotic LcS is highly sensitive to acidic conditions and does not show viability when immersed in the stomach under fasting conditions.27 The greatest extent of aflatoxin binding by LcS was at pH 7.2.24 Thus, it is strongly advised to consume probiotics after meals to maximise the beneficial effects.

Consuming a high intake of carbohydrates, fats, and/or proteins can trigger the secretion of digestive juices,28,29 which may not be adapted by all probiotics. However, a recent in vivo study by Nurul Adilah et al. (2018) did not agree as the authors demonstrated a protective effect against urinary AFM1 levels in rats who received LcS supplementation with a high protein diet.15 These findings lend for further studies to assess their interaction.

Probiotics may be less capable of binding aflatoxin in the presence of intestinal mucus.20 Despite that, their binding sites on probiotics were unlikely to be similar as AFB1 is known to bind to carbohydrates25 while mucus must adhere to proteins.30 The addition of proteins to mucus in the intestinal tract may minimise the reductive effects of AFB1 binding by probiotics.20 It also leads to the question of whether dietary fibre could mask the probiotic binding efficacy since dietary fibre is responsible for mucus production.31 The mechanisms of the interaction between probiotics, aflatoxins, and intestinal mucus deserved further investigation.


Probiotic consumption may be a practical dietary approach to prevent human exposure to aflatoxins. Future trials are warranted to explore the factors affecting probiotic efficacy as they may explain the conflict of results observed between in vitro and in vivo with human studies.


  1. Zheng, J., Wittouck, S., Salvetti, E., Franz, C. M., Harris, H. M., Mattarelli, P., O’Toole, P. W., Pot, B., Vandamme, P., & Walter, J. (2020). A taxonomic note on the genus Lactobacillus: Description of 23 novel genera, emended description of the genus Lactobacillus Beijerinck 1901, and union of Lactobacillaceae and Leuconostocaceae. International Journal of Systematic and Evolutionary Microbiology, 70(4), 2782-2858.
  2. Tang, X., & Zhao, J. (2019). Commercial strains of lactic acid bacteria with health benefits. In W. Chen (Ed.), Lactic Acid Bacteria (pp. 297-369). Springer.
  3. Siti Soleha, A. D., Aida Nurul Ain, A. S., Nur Azra, M. P., Hasiah, A. H., Mohd Redzwan, S., & Rozaini, A. (2022). Aflatoxin B1 reported in herbal and traditional medicine and its risk assessment. Malaysian Journal of Medicine and Health Sciences, 18, 31-37.
  4. Costamagna, D., Gaggiotti, M., Chiericatti, C. A., Costabel, L., Audero, G. M. d. L., Taverna, M., & Signorini, M. L. (2019). Quantification of aflatoxin M1 carry-over rate from feed to soft cheese. Toxicology Reports, 6, 782-787.
  5. Farah Nadira, A., Rosita, J., Norhaizan, M. E., & Mohd Redzwan, S. (2017). Screening of aflatoxin M1 occurrence in selected milk and dairy products in Terengganu, Malaysia. Food Control, 73, 209-214.
  6. Dhakal, A., & Sbar, E. (2022, May 23, 2022). Aflatoxin Toxicity. StatPearls Publishing. Retrieved 8 Aug, 2022 from https://www.ncbi.nlm.nih.gov/books/NBK557781/
  7. Nurul Adilah, Z., & Mohd Redzwan, S. (2017). Effect of dietary macronutrients on aflatoxicosis: A mini-review. Journal of the Science of Food and Agriculture, 97(8), 2277-2281.
  8. International Agency for Research on Cancer. (2022). Agents Classified by the IARC Monographs. Volumes 1-132. https://monographs.iarc.who.int/agents-classified-by-the-iarc/
  9. World Health Organization. (2017). Evaluation of certain contaminants in food. Eighty-third report of the Joint FAO/WHO Expert Committee on Food Additives. World Health Organization. https://apps.who.int/iris/bitstream/handle/10665/254893/9789241210027-eng.pdf
  10. Liew, W. P. P., Mohd Redzwan, S., Than, L. T. L., & Fauzah, A. G. (2022). Metagenomic and proteomic approaches in elucidating aflatoxin B1 detoxification mechanisms of probiotic Lactobacillus casei Shirota towards intestine. Food and Chemical Toxicology, 160, 112808-112808.
  11. Mohd Redzwan, S., Rosita, J., Farah Nadira, A., & Lim, Y. J. (2016). Chapter 27. Probiotics as potential adsorbent of aflatoxin. In W. Ronald Ross & R. P. Victor (Eds.), robiotics, Prebiotics, and Synbiotics. (pp. 409-419). Elsevier.
  12. Liew, W. P. P., Nurul Adilah, Z., Than, L. T. L., & Mohd Redzwan, S. (2018). The binding efficiency and interaction of Lactobacillus casei Shirota toward aflatoxin B1. Frontiers in Microbiology, 9, 1503.
  13. Nikbakht, E., Jamaluddin, R., Abdul Mutalib, M. S., Khaza’ai, H., Khalesi, S., & Mohd Redzwan, S. (2013). Reduction of aflatoxin level in aflatoxin-induced rats by the activity of probiotic Lactobacillus casei strain Shirota. Journal of Applied Microbiology, 114(5), 1507-1515.
  14. Nikbakht, E., Jamaluddin, R., Redzwan, S. M., & Khalesi, S. (2019). Oral administration of Lactobacillus casei Shirota can ameliorate the adverse effect of an acute aflatoxin exposure in Sprague Dawley rats. International Journal for Vitamin and Nutrition Research, 88(3-4), 199-208.
  15. Nurul Adilah, Z., Liew, W. P. P., Mohd Redzwan, S., & Amin, I. (2018). Effect of high protein diet and probiotic Lactobacillus casei Shirota supplementation in aflatoxin B1-induced rats. BioMed Research International, 2018.
  16. Kumar, M., Verma, V., Nagpal, R., Kumar, A., Behare, P. V., Singh, B., & Aggarwal, P. K. (2012). Anticarcinogenic effect of probiotic fermented milk and chlorophyllin on aflatoxin-B1-induced liver carcinogenesis in rats. British Journal of Nutrition, 107(7), 1006-1016.
  17. Hernandez Mendoza, A., Guzman De Peña, D., González Córdova, A. F., Vallejo Córdoba, B., & Garcia, H. S. (2010). in vivo assessment of the potential protective effect of Lactobacillus casei Shirota against aflatoxin B1. Dairy Science and Technology, 90(6), 729-740.
  18. Cox, A. J., Makino, H., Cripps, A. W., & West, N. P. (2019). Recovery of Lactobacillus casei strain Shirota (LcS) from faeces with 14 days of fermented milk supplementation in healthy Australian adults. Asia Pacific Journal of Clinical Nutrition, 28(4), 734.
  19. Wang, C., Nagata, S., Asahara, T., Yuki, N., Matsuda, K., Tsuji, H., Takahashi, T., Nomoto, K., & Yamashiro, Y. (2015). Intestinal microbiota profiles of healthy pre-school and school-age children and effects of probiotic supplementation. Annals of Nutrition and Metabolism, 67(4), 257-266.
  20. Gratz, S., Mykkänen, H., Ouwehand, A. C., Juvonen, R., Salminen, S., & El-Nezami, H. (2004). Intestinal mucus alters the ability of probiotic bacteria to bind aflatoxin B1 in vitro. Applied and Environmental Microbiology, 70(10), 6306-6308.
  21. Mohd Redzwan, S., Mohd Sokhini, A. M., Wang, J. S., Ahmad, Z., Kang, M. S., Nasrabadi, E. N., & Rosita, J. (2016). Effect of supplementation of fermented milk drink containing probiotic Lactobacillus casei Shirota on the concentrations of aflatoxin biomarkers among employees of Universiti Putra Malaysia: A randomised, double-blind, cross-over, placebo-controlled study. British Journal of Nutrition, 115(1), 39-54.
  22. Liew, W. P. P., Mohd Redzwan, S., & Than, L. T. L. (2019). Gut microbiota profiling of aflatoxin B1-induced rats treated with Lactobacillus casei Shirota. Toxins, 11(1), 49.
  23. Afshar, P., Shokrzadeh, M., Raeisi, S. N., Ghorbani-HasanSaraei, A., & Nasiraii, L. R. (2020). Aflatoxins biodetoxification strategies based on probiotic bacteria. Toxicon, 178, 50-58.
  24. Hernandez Mendoza, A., Guzman de Peña, D., & Garcia, H. S. (2009). Key role of teichoic acids on aflatoxin B1 binding by probiotic bacteria. Journal of Applied Microbiology, 107(2), 395-403.
  25. Haskard, C., Binnion, C., & Ahokas, J. (2000). Factors affecting the sequestration of aflatoxin by Lactobacillus rhamnosus strain GG. Chemico-Biological Interactions, 128(1), 39-49.
  26. El-Nezami, H., Kankaanpaa, P., Salminen, S., & Ahokas, J. (1998). Ability of dairy strains of lactic acid bacteria to bind a common food carcinogen, aflatoxin B1. Food and Chemical Toxicology, 36(4), 321-326.
  27. Caillard, R., & Lapointe, N. (2017). In vitro gastric survival of commercially available probiotic strains and oral dosage forms. International Journal of Pharmaceutics, 519(1-2), 125-127.
  28. Boivin, M., Lanspa, S. J., Zinsmeister, A. R., Go, V. L. W., & DiMagno, E. P. (1990). Are diets associated with different rates of human interdigestive and postprandial pancreatic enzyme secretion? Gastroenterology, 99(6), 1763-1771.
  29. Hara, H., Ochi, Y., & Kasai, T. (1998). Bile-pancreatic juice-independent increases in pancreatic proteases and intestinal cholecystokinin by dietary protein in rats. Proceedings of the Society for Experimental Biology and Medicine, 217(2), 173-179.
  30. Tuomola, E. M., Ouwehand, A. C., & Salminen, S. J. (2000). Chemical, physical and enzymatic pre-treatments of probiotic lactobacilli alter their adhesion to human intestinal mucus glycoproteins. International Journal of Food Microbiology, 60(1), 75-81.
  31. Brownlee, I. A., Havler, M. E., Dettmar, P. W., Allen, A., & Pearson, J. P. (2003). Colonic mucus: Secretion and turnover in relation to dietary fibre intake. Proceedings of the Nutrition Society, 62(1), 245-249.
About the Authors

Ms. Chang Wei Lin holds a MSc degree in Nutritional Science at the Universiti Putra Malaysia. She is studying her Ph.D under the main supervision of Dr. Mohd Redzwan Sabran. She is engaged in clinical trials, exploring the effects of probiotics in reducing aflatoxins.

Dr. Mohd Redzwan Sabran is a senior lecturer in Nutrition at the Universiti Putra Malaysia. Dr. Redzwan has been engaged in teaching and research for more than 7 years. His expertise includes Nutritional Sciences, Food Safety and Food Contaminants, as well as Probiotics.

Both authors are from Department of Nutrition, Faculty of Medicine and Health Sciences, University Putra Malaysia

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