Metabolomic approach to evaluating the effectiveness of Lactobacillus reuteri LR92 probiotic treatment for infantile colic in breastfed children

Introduction. Symptoms typically peak around the sixth week of life and resolve spontaneously by 4–6 months of age, which is why colic is generally considered a benign and self-limiting condition. The need for treatment arises from the negative impact of excessive crying on the infant’s family, as well as the potential long-term consequences of colic. The effectiveness of Lactobacillus reuteri DSM 17938 in breastfed infants has been demonstrated. Recently, there has been growing interest in the LR92 strain; however, its mechanism of action remains unclear.

Aim. To study the urine metabolome in 1–4 month-old breastfed infants receiving Lactobacillus reuteri LR92 probiotic (Maxilac^® Baby drops) for the treatment of infantile colic, identify probable mechanisms of action of this probiotic, and assess its safety.

Materials and methods. A clinical post-registration open observational prospective single-center study with minimal intervention and a comparison group was conducted: 38 children aged 1 to 5 months who were breastfed and diagnosed with infantile colic. Patients were divided into two groups: the main group (17 children) received Maxilac^® Baby probiotic drops for oral administration, the comparison group (21 children) did not receive probiotics. Urine metabolome analysis was performed using gas chromatography-mass spectrometry (GC-MS) with modern chromatographic equipment.

Results and discussion. Children with colic showed significant changes in urine metabolome, reflecting impaired intestinal absorption, energy metabolism, and microbiocenosis. Following treatment with probiotic, complete resolution of colic by day 14 was observed in 100% of children in the main group (compared to only 20% in the comparison group); decreased urine content of short-chain fatty acids; changes in neurotransmitter levels indicating suppression of excitatory and stimulation of inhibitory neurotransmitters; absence of side effects.

Conclusions. Maxilac^® Baby probiotic demonstrated high efficacy and safety in treating infantile colic, confirmed by clinical and metabolomic studies. The mechanism of action is associated with normalization of microbiocenosis, metabolic processes, and neurotransmitter status.

1. Steutel NF, Zeevenhooven J, Scarpato E, Vandenplas Y, Tabbers MM, Staiano A et al. Prevalence of functional gastrointestinal disorders in european infants and toddlers. J Pediatr. 2020;221:107–114. https://doi.org/10.1016/j.jpeds.2020.02.076.

2. Wolke D, Bilgin A, Samara M. Systematic review and meta-analysis: fussing and crying durations and prevalence of colic in infants. J Pediatr. 2017;185:55–61. https://doi.org/10.1016/j.jpeds.2017.02.020.

3. Benninga MA, Faure C, Hyman PE, St James Roberts I, Schechter NL, Nurko S. Childhood functional gastrointestinal disorders: neonate/toddler. Gastroenterology. 2016;150(6):1443–1455. https://doi.org/10.1053/j.gastro.2016.02.016.

4. Gurova ММ. Infantile colic from the perspective of evidence-based medicine: prevalence, principles of differential diagnosis and diet therapy algorithm. Meditsinskiy Sovet. 2019;(17):147–155. (In Russ.) https://doi.org/10.21518/2079-701X-2019-17-147-155.

5. Drossman DA. Functional gastrointestinal disorders: history, pathophysiology, clinical features, and Rome IV. Gastroenterology. 2016;150:1262–1279e2. https://doi.org/10.1053/j.gastro.2016.02.032.

6. Novikova VP, Magamedova DM. Infantile colic. Children’s Medicine of the North-West. 2024;12(1):29–46. (In Russ.) https://doi.org/10.56871/CmN-W.2024.42.28.003.

7. Khavkin AI, Magamedova DM, Novikova VP. Infantile colic: facets of the problem. Pediatric Nutrition. 2024;22(2):60–73. (In Russ.) https://doi.org/10.20953/1727-5784-2024-2-60-72.

8. Hofman D, Kudla U, Miqdady M, Nguyen TVH, Morán-Ramos S, Vandenplas Y. Faecal microbiota in infants and young children with functional gastrointestinal disorders: a systematic review. Nutrients. 2022;14(5):974. https://doi.org/10.3390/nu14050974.

9. Kozhakhmetov S, Meiirmanova Z, Mukhanbetzhanov N, Jarmukhanov Z, Vinogradova E, Mureyev S et al. Compositional and functional variability of the gut microbiome in children with infantile colic. Sci Rep. 2023;13(1):9530. https://doi.org/10.1038/s41598-023-36641-z.

10. Turunen J, Tejesvi MV, Paalanne NP, Pokka T, Amatya SB, Mishra S et al. Investigating prenatal and perinatal factors on meconium microbiota: a systematic review and cohort study. Pediatr Res. 2024;95(1):135–145. https://doi.org/10.1038/s41390-023-02783-z.

11. Rosa D, Zablah RA, Vazquez-Frias R. Unraveling the complexity of Disorders of the Gut-Brain Interaction: the gut microbiota connection in children. Front Pediatr. 2024;11:1283389. https://doi.org/10.3389/fped.2023.1283389.

12. O’Mahony SM, Clarke G, Borre YE, Dinan TG, Cryan JF. Serotonin, Tryptophan Metabolism and the Brain-Gut-Microbiome Axis. Behav Brain Res. 2015;277:32–48. https://doi.org/10.1016/j.bbr.2014.07.027.

13. Hoyles L, Snelling T, Umlai UK, Nicholson JK, Carding SR, Glen RC, McArthur S. Microbiome-Host Systems Interactions: Protective Effects of Propionate upon the Blood-Brain Barrier. Microbiome. 2018;6:55. https://doi.org/10.1186/s40168-018-0439-y.

14. Dalile B, Van Oudenhove L, Vervliet B, Verbeke K. The role of short-chain fatty acids in microbiota-gut-brain communication. Nat Rev Gastroenterol Hepatol. 2019;16(8):461–478. https://doi.org/10.1038/s41575-019-0157-3.

15. Cryan JF, O’Riordan KJ, Cowan CSM, Sandhu KV, Bastiaanssen TFS, Boehme M et al. The microbiota-gut-brainaxis. Physiol Rev. 2019;99(4):1877–2013. https://doi.org/10.1152/physrev.00018.2018.

16. Silva YP, Bernardi A, Frozza RL. The Role of Short-Chain Fatty Acids From Gut Microbiota in Gut-Brain Communication. Front Endocrinol. 2020;11:25. https://doi.org/10.3389/fendo.2020.00025.

17. Khavkin AI, Bogdanova NM, Belova EM. Role of neuropeptides in the genesis of intestinal colic. Farmateka. 2019;26(2):89–92. (In Russ.) https://doi.org/10.18565/pharmateca.2019.2.89-92.

18. McNabney SM, Henagan TM. Short Chain Fatty Acids in the Colon and Peripheral Tissues: Role in Health and Disease. Nutrients. 2017;9(12):1348. https://doi.org/10.3390/nu9121348.

19. Koh A, De Vadder F, Kovatcheva-Datchary P, Bäckhed F. From Dietary Fiber to Host Physiology: Short-Chain Fatty Acids as Key Bacterial Metabolites. Cell. 2016;165(6):1332–1345. https://doi.org/10.1016/j.cell.2016.05.041.

20. Ríos-Covián D, Ruas-Madiedo P, Margolles A, Gueimonde M, de Los Reyes-Gavilán CG, Salazar N et al. Intestinal Short Chain Fatty Acids and their Link with Diet and Human Health. Front Microbiol. 2016;7:185. https://doi.org/10.3389/fmicb.2016.00185.

21. Dalile B, Van Oudenhove L, Vervliet B, Verbeke K. The Role of Short-Chain Fatty Acids in Microbiota–Gut–Brain Communication. Nat Rev Gastroenterol Hepatol. 2019;16(8):461–478. https://doi.org/10.1038/s41575-019-0157-3.

22. Frost G, Sleeth ML, Sahuri-Arisoylu M, Lizarbe B, Cerdan S, Brody L et al. The short-chain fatty acid acetate reduces appetite via a central homeostatic mechanism. Nat Commun. 2014;5:3611. https://doi.org/10.1038/ncomms4611.

23. de Weerth C. Do bacteria shape our development? Crosstalk between intestinal microbiota and HPA axis. Neurosci Biobehav Rev. 2017;83:458–471. https://doi.org/10.1016/j.neubiorev.2017.09.016.

24. O’Riordan KJ, Collins MK, Moloney GM, Knox EG, Aburto MR, Fülling C et al. Short chain fatty acids: Microbial metabolites for gut-brain axis signalling. Mol Cell Endocrinol. 2022;546:111572. https://doi.org/10.1016/j.mce.2022.111572.

25. Peng Y, Ma Y, Luo Z, Jiang Y, Xu Z, Yu R. Lactobacillus reuteri in digestive system diseases: focus on clinical trials and mechanisms. Front Cell Infect Microbiol. 2023;13:1254198. https://doi.org/10.3389/fcimb.2023.1254198.

26. Chen Z, Kim J. Urinary proteomics and metabolomics studies to monitor bladder health and urological diseases. BMC Urol. 2016;16:11. https://doi.org/10.1186/s12894-016-0129-7.

27. Novikova, VN, Magamedova, DM. Probiotic properties of lactobacillus reuteri (L. Reuteri) strains. Children’s Medicine of the North-West. 2023;11(3):36–53. (In Russ.) https://doi.org/10.56871/CmN-W.2023.75.34.002.

28. Pourmirzaiee MA, Famouri F, Moazeni W, Hassanzadeh A, Hajihashemi M. The efficacy of the prenatal administration of Lactobacillus reuteri LR92 DSM 26866 on the prevention of infantile colic: a randomized control trial. Eur J Pediatr. 2020;179(10):1619–1626. https://doi.org/10.1007/s00431-020-03641-4.

29. Savino F, Pelle E, Palumeri E, Oggero R, Miniero R. Lactobacillus reuteri (American Type Culture Collection Strain 55730) Versus Simethicone in the Treatment of Infantile Colic: A Prospective Randomized Study. Pediatrics. 2007;119:e124–e130. https://doi.org/10.1542/peds.2006-1222.

30. Dryl R, Szajewska H. Probiotics for management of infantile colic: a systematic review of randomized controlled trials. Arch Med Sci. 2018;14(5):1137–1143. https://doi.org/10.5114/aoms.2017.66055.

31. Savino F, Garro M, Montanari P, Galliano I, Bergallo M. Crying Time and RORγ/FOXP3 Expression in Lactobacillus reuteri DSM17938-Treated Infants with Colic: A Randomized Trial. J Pediatr. 2018;192:171–177.e1. https://doi.org/10.1016/j.jpeds.2017.08.062.

32. Hojsak I. Probiotics in functional gastrointestinal disorders. Adv Exp Med Biol. 2019;1125:121–137. https://doi.org/10.1007/5584_2018_321.

33. Ivashkin VT, Gorelov AV, Abdulganieva DI, Alekseeva OP, Alekseenko SA, Baranovsky AYu et al. Methodological Guidelines of the Scientific Community for Human Microbiome Research (CHMR) and the Russian Gastroenterology Association (RGA) on the Use of Probiotics, Prebiotics, Synbiotics, Metabiotics and Functional Foods Enriched with Them for the Treatment and Prevention of Gastrointestinal Diseases in Adults and Children. Russian Journal of Gastroenterology, Hepatology, Coloproctology. 2024;34(4):113–136. (In Russ.) https://doi.org/10.22416/1382-4376-2024-117-312.

34. Vandenplas Y, Hauser B, Salvatore S. Functional Gastrointestinal Disorders in Infancy: Impact on the Health of the Infant and Family. Pediatr Gastroenterol Hepatol Nutr. 2019;22:207. https://doi.org/10.5223/pghn.2019.22.3.207.

35. Zhen J, Zhou Z, He M, Han HX, Lv EH, Wen PB et al. The gut microbial metabolite trimethylamine N-oxide and cardiovascular diseases. Front Endocrinol. 2023;14:1085041. https://doi.org/10.3389/fendo.2023.1085041.

36. Al-Obaide MAI, Singh R, Datta P, Rewers-Felkins KA, Salguero MV, Al-Obaidi I et al. Gut Microbiota-Dependent Trimethylamine-N-oxide and Serum Biomarkers in Patients with T2DM and Advanced CKD. J Clin Med. 2017;6(9):86. https://doi.org/10.3390/jcm6090086.

37. Hoyles L, Snelling T, Umlai UK, Nicholson JK, Carding SR, Glen RC, McArthur S. Microbiome-host systems interactions: protective effects of propionate upon the blood-brain barrier. Microbiome. 2018;6(1):55. https://doi.org/10.1186/s40168-018-0439-y.

38. Yuan S, Jin Z, Ali A, Wang C, Liu J. Caproic Acid-Producing Bacteria in Chinese Baijiu Brewing. Front Microbiol. 2022;13:883142. https://doi.org/10.3389/fmicb.2022.883142.

39. Xiong RG, Zhou DD, Wu SX, Huang SY, Saimaiti A, Yang ZJ et al. Health Benefits and Side Effects of Short-Chain Fatty Acids. Foods. 2022;11(18):2863. https://doi.org/10.3390/foods11182863.

40. Fusco W, Lorenzo MB, Cintoni M, Porcari S, Rinninella E, Kaitsas F et al. Short-Chain Fatty-Acid-Producing Bacteria: Key Components of the Human Gut Microbiota. Nutrients. 2023;15(9):2211. https://doi.org/10.3390/nu15092211.

41. Grüter T, Nuwin M, Rilke N, Pitarokoili K. Propionate exerts neuroprotective and neuroregenerative effects in the peripheral nervous system. Proc Natl Acad Sci U S A. 2023;120(4):e2216941120. https://doi.org/10.1073/pnas.2216941120.

42. Blaak EE, Canfora EE, Theis S, Frost G, Groen AK, Mithieux G et al. Short chain fatty acids in human gut and metabolic health. Benef Microbes. 2020;11(5):411–455. https://doi.org/10.3920/BM2020.0057.

43. Torshin IIu, Gromova OA, Zgoda VG, Tikhonova OV, Malyavskaya SI. Cerebrolysin peptides as mood stabilizers. Zhurnal Nevrologii i Psikhiatriiimeni S.S. Korsakova. 2019;119(12):69–75. (In Russ.) https://doi.org/10.17116/jnevro201911912169.

44. Fragkos KC, Forbes A. Citrulline as a marker of intestinal function and absorption in clinical settings: A systematic review and metaanalysis. United European Gastroenterol J. 2018;(6):181–191. https://doi.org/10.1177/2050640617737632.

45. Khavkin AI, Novikova VP, Shapovalova NS. Promising non-invasive biomarkers: intestinal proteins in the diagnosis of intestinal mucosal damage. Experimental and Clinical Gastroenterology. 2021;188(4):155–160. (In Russ.) https://doi.org/10.31146/1682-8658-ecg-188-4-155-160.

46. Averyanova NI, Balueva LG, Ivanova NV, Rudavina TI. Disorder of oxalic acid metabolism in children. Modern Problems of Science and Education. 2015;(3). (In Russ.) Available at: https://science-education.ru/ru/article/view?id=19738.

47. Sitkin SI, Vakhitov TYa, Tkachenko EI, Lazebnik LB, Oreshko LS, Zhigalova TN et al. Gut microbial and endogenous metabolism alterations in ulcerative colitis and celiac disease: a metabolomics approach to identify candidate biomarkers of chronic intestinal inflammation associated with dysbiosis. Experimental and Clinical Gastroenterology. 2017;(7):4–50. (In Russ.) Available at: https://www.nogr.org/jour/article/view/449.