ANALYSIS OF THE EFFECTS OF MAGNESIUM CITRATE AND PROTEINOGENIC BRANCHED-CHAIN AMINO ACIDS ON THE CONCENTRATION OF CALCIPROTEIN PARTICLES IN CIRCULATING BLOOD

Highlights

• Oral administration of proteinogenic branched-chain amino acids (leucine, isoleucine, and valine) and magnesium citrate to hyperlipidemic mice did not affect the concentration of calciprotein particles in the serum.
• The addition of magnesium citrate to calcium- and phosphate-supersaturated serum obtained from patients with ST-segment elevation myocardial infarction resulted in a statistically significant decrease in calciprotein particle concentration.
• These findings suggest the potential utility of magnesium citrate for reducing the circulation of calciprotein particles in the blood as soon as they aggregate excessive calcium and phosphate.

Aim. To evaluate the effects of proteinogenic branched-chain amino acids (leucine, isoleucine, and valine; BCAA) and magnesium citrate on the concentration of calciprotein particles (CPPs) in the blood of hyperlipidemic (ApoE-knockout) mice, as well as in the serum of patients with ST-segment elevation myocardial infarction (STEMI) following ex vivo addition of magnesium citrate.

Methods. The study included 270 ApoE-knockout mice (135 male and 135 female mice) that received BCAA (55 mg/day, equivalent to a human equivalent dose of 13.5 g/day) or magnesium citrate (1.85 mg/day, equivalent to a human equivalent dose of 450 mg/day), starting from the age of 1, 3, or 6 months (90 mice per group; 45 male and 45 female mice in each). At 12 months of age, all animals were euthanized, and serum CPP concentrations were quantified using flow cytometry with a fluorescently labeled bisphosphonate probe. Additionally, serum samples from 27 patients with STEMI were analyzed ex vivo by inducing mineral stress through the addition of CaCl₂ and Na₂HPO₄·12H₂O, followed by magnesium citrate supplementation to assess its capacity to attenuate CPP formation.

Results. Oral administration of BCAA or magnesium citrate in ApoE-knockout mice did not affect serum CPP concentrations. In contrast, the addition of magnesium citrate to calcium- and phosphate-supersaturated serum from STEMI patients led to a reduction in CPP concentration in 74% of cases (20/27), with an over 81% (22/27) rate of reduction compared to the respective non-supersaturated serum.

Conclusion. The obtained data suggest a potential therapeutic effect of magnesium citrate, which may exert chelating properties and promote the dissolution of circulating CPPs in the blood.

1. Nielsen RV, Fuster V, Bundgaard H, Fuster JJ, Johri AM, Kofoed KF, Douglas PS, Diederichsen A, Shapiro MD, Nicholls SJ, Nordestgaard BG, Lindholt JS, MacRae C, Yu-an C, Newby DE, Urbina EM, Bergström G, Ridderstråle M, Budoff MJ, Bøttcher M, Rai-takari OT, Hansen TH, Näslund U, Sillesen H, Eldrup N, Ibanez B. Personalized Interven-tion Based on Early Detection of Atherosclerosis: JACC State-of-the-Art Review. J Am Coll Cardiol. 2024 May 28;83(21):2112-2127. doi: 10.1016/j.jacc.2024.02.053..

2. McPherson R, Adreak N, Sharma A. Medications for Lipid Control: Statins vs New-er Drugs. Can J Cardiol. 2024 Aug;40(8S):S26-S34. doi: 10.1016/j.cjca.2024.05.004.

3. Tokgözoğlu L, Libby P. The dawn of a new era of targeted lipid-lowering therapies. Eur Heart J. 2022;43(34):3198-3208. doi: 10.1093/eurheartj/ehab841.

4. Global Burden of Cardiovascular Diseases and Risks 2023 Collaborators. Global, Regional, and National Burden of Cardiovascular Diseases and Risk Factors in 204 Coun-tries and Territories, 1990-2023. J Am Coll Cardiol. 2025 Sep 24:S0735-1097(25)07428-5. doi: 10.1016/j.jacc.2025.08.015.

5. Karimian J, Abedi S, Shirinbakhshmasoleh M, Moodi F, Moodi V, Ghavami A. The effects of quinoa seed supplementation on cardiovascular risk factors: A systematic review and meta-analysis of controlled clinical trials. Phytother Res. 2021;35(4):1688-1696. doi: 10.1002/ptr.6901.

6. Banach M, Patti AM, Giglio RV, Cicero AFG, Atanasov AG, Bajraktari G, Bruckert E, Descamps O, Djuric DM, Ezhov M, Fras Z, von Haehling S, Katsiki N, Langlois M, Lat-kovskis G, Mancini GBJ, Mikhailidis DP, Mitchenko O, Moriarty PM, Muntner P, Nikolic D, Panagiotakos DB, Paragh G, Paulweber B, Pella D, Pitsavos C, Reiner Ž, Rosano GMC, Rosenson RS, Rysz J, Sahebkar A, Serban MC, Vinereanu D, Vrablík M, Watts GF, Wong ND, Rizzo M; International Lipid Expert Panel (ILEP). The Role of Nutraceuticals in Statin Intolerant Patients. J Am Coll Cardiol. 2018;72(1):96-118. doi: 10.1016/j.jacc.2018.04.040.

7. Riccardi G, Giosuè A, Calabrese I, Vaccaro O. Dietary recommendations for preven-tion of atherosclerosis. Cardiovasc Res. 2022;118(5):1188-1204. doi: 10.1093/cvr/cvab173.

8. Li Z, Zhang R, Mu H, Zhang W, Zeng J, Li H, Wang S, Zhao X, Chen W, Dong J, Yang R. Oral Administration of Branched-Chain Amino Acids Attenuates Atherosclerosis by Inhibiting the Inflammatory Response and Regulating the Gut Microbiota in ApoE-Deficient Mice. Nutrients. 2022;14(23):5065. doi: 10.3390/nu14235065.

9. Lee JH, Park E, Jin HJ, Lee Y, Choi SJ, Lee GW, Chang PS, Paik HD. Anti-inflammatory and anti-genotoxic activity of branched chain amino acids (BCAA) in lipo-polysaccharide (LPS) stimulated RAW 264.7 macrophages. Food Sci Biotechnol. 2017;26(5):1371-1377. doi: 10.1007/s10068-017-0165-4.

10. Ijichi C, Matsumura T, Tsuji T, Eto Y. Branched-chain amino acids promote albumin synthesis in rat primary hepatocytes through the mTOR signal transduction system. Bio-chem Biophys Res Commun. 2003;303(1):59-64. doi: 10.1016/s0006-291x(03)00295-x.

11. Ichikawa K, Okabayashi T, Shima Y, Iiyama T, Takezaki Y, Munekage M, Namika-wa T, Sugimoto T, Kobayashi M, Mimura T, Hanazaki K. Branched-chain amino acid-enriched nutrients stimulate antioxidant DNA repair in a rat model of liver injury induced by carbon tetrachloride. Mol Biol Rep. 2012;39(12):10803-10. doi: 10.1007/s11033-012-1974-4.

12. Hsu YM, Kuan HC, Chen YA, Chiu CW, Chen PC, Tam KW. Effects of branched-chain amino acids supplementation on patients undergoing hepatic intervention: a meta-analysis of randomised controlled trials. Br J Nutr. 2024;131(2):276-285. doi: 10.1017/S0007114523001885.

13. Zoanni B, Brioschi M, Mallia A, Gianazza E, Eligini S, Carini M, Aldini G, Banfi C. Novel insights about albumin in cardiovascular diseases: Focus on heart failure. Mass Spec-trom Rev. 2023;42(4):1113-1128. doi: 10.1002/mas.21743.

14. Heiss A, Eckert T, Aretz A, Richtering W, van Dorp W, Schäfer C, Jahnen-Dechent W. Hierarchical role of fetuin-A and acidic serum proteins in the formation and stabilization of calcium phosphate particles. J Biol Chem. 2008;283(21):14815-25. doi: 10.1074/jbc.M709938200.

15. Taverna M, Marie AL, Mira JP, Guidet B. Specific antioxidant properties of human serum albumin. Ann Intensive Care. 2013;3(1):4. doi: 10.1186/2110-5820-3-4.

16. Rennie C, Donnelly S, McGrath K. Albumin Reduces Hepatic Steatosis and Inflam-mation in High-Fat-Diet-Fed Mice. Int J Mol Sci. 2025;26(15):7156. doi: 10.3390/ijms26157156.

17. Paar M, Rossmann C, Nusshold C, Wagner T, Schlagenhauf A, Leschnik B, Oettl K, Koestenberger M, Cvirn G, Hallström S. Anticoagulant action of low, physiologic, and high albumin levels in whole blood. PLoS One. 2017;12(8):e0182997. doi: 10.1371/journal.pone.0182997.

18. Seidu S, Kunutsor SK, Khunti K. Serum albumin, cardiometabolic and other adverse outcomes: systematic review and meta-analyses of 48 published observational cohort studies involving 1,492,237 participants. Scand Cardiovasc J. 2020;54(5):280-293. doi: 10.1080/14017431.2020.1762918.

19. Thuemmler RJ, Pana TA, Carter B, Mahmood R, Bettencourt-Silva JH, Metcalf AK, Mamas MA, Potter JF, Myint PK. Serum Albumin and Post-Stroke Outcomes: Analysis of UK Regional Registry Data, Systematic Review, and Meta-Analysis. Nutrients. 2024;16(10):1486. doi: 10.3390/nu16101486.

20. Yao Z, Xu Y, Ma W, Sun XY, Jia S, Zheng Y, Liu X, Fan Y, Wang C. Magnesium Citrate Protects Against Vascular Calcification in an Adenine-induced Chronic Renal Fail-ure Rat Model. J Cardiovasc Pharmacol. 2018;72(6):270-276. doi: 10.1097/FJC.0000000000000590.

21. Quiñones H, Hamdi T, Sakhaee K, Pasch A, Moe OW, Pak CYC. Control of meta-bolic predisposition to cardiovascular complications of chronic kidney disease by efferves-cent calcium magnesium citrate: a feasibility study. J Nephrol. 2019;32(1):93-100. doi: 10.1007/s40620-018-0559-2.

22. Zanforlini BM, Ceolin C, Trevisan C, Alessi A, Seccia DM, Noale M, Maggi S, Guarnieri G, Vianello A, Sergi G. Clinical trial on the effects of oral magnesium supplemen-tation in stable-phase COPD patients. Aging Clin Exp Res. 2022;34(1):167-174. doi: 10.1007/s40520-021-01921-z.

23. Deng K, Liu J, Miao Y, Wang G, Wang X, Liu S, Yang L. The effects of magnesium and vitamin D/E co-supplementation on inflammation markers and lipid metabolism of obese/overweight population: a systematic review and meta-analysis. Front Nutr. 2025;12:1563604. doi: 10.3389/fnut.2025.1563604.

24. Darooghegi Mofrad M, Djafarian K, Mozaffari H, Shab-Bidar S. Effect of magnesi-um supplementation on endothelial function: A systematic review and meta-analysis of ran-domized controlled trials. Atherosclerosis. 2018;273:98-105. doi: 10.1016/j.atherosclerosis.2018.04.020.

25. Argeros Z, Xu X, Bhandari B, Harris K, Touyz RM, Schutte AE. Magnesium Sup-plementation and Blood Pressure: A Systematic Review and Meta-Analysis of Randomized Controlled Trials. Hypertension. 2025;82(11):1844-1856. doi: 10.1161/HYPERTENSIONAHA.125.25129.

26. Verma H, Garg R. Effect of magnesium supplementation on type 2 diabetes associat-ed cardiovascular risk factors: a systematic review and meta-analysis. J Hum Nutr Diet. 2017;30(5):621-633. doi: 10.1111/jhn.12454.

27. Hariri M, Sohrabi M, Gholami A. The effect of magnesium supplementation on serum concentration of lipid profile: an updated systematic review and dose-response meta-analysis on randomized controlled trials. Nutr J. 2025;24(1):24. doi: 10.1186/s12937-025-01085-w.

28. Liu H, Wang R. Associations between the serum magnesium and all-cause or cardio-vascular mortality in chronic kidney disease and end-stage renal disease patients: A meta-analysis. Medicine (Baltimore). 2021;100(45):e27486. doi: 10.1097/MD.0000000000027486.

29. Kobylecki CJ, Nordestgaard BG, Afzal S. Plasma Ionized Calcium and Risk of Car-diovascular Disease: 106 774 Individuals from the Copenhagen General Population Study. Clin Chem. 2021;67(1):265-275. doi: 10.1093/clinchem/hvaa245.

30. Kobylecki CJ, Nordestgaard BG, Afzal S. Low Plasma Ionized Calcium Is Associat-ed With Increased Mortality: A Population-based Study of 106 768 Individuals. J Clin En-docrinol Metab. 2022;107(7):e3039-e3047. doi: 10.1210/clinem/dgac146.

31. Mora S, Mann G, Adegoke OAJ. Sex differences in cachexia and branched-chain amino acid metabolism following chemotherapy in mice. Physiol Rep. 2024;12(8):e16003. doi: 10.14814/phy2.16003.

32. Liu K, Borreggine R, Gallart-Ayala H, Ivanisevic J, Marques-Vidal P. Serum branched-chain amino acids are mainly associated with body mass index and waist circum-ference. Nutr Metab Cardiovasc Dis. 2025;35(7):103880. doi: 10.1016/j.numecd.2025.103880.

33. Muscella A, Felline M, Marsigliante S. Sex-Based Effects of Branched-Chain Amino Acids on Strength Training Performance and Body Composition. Sports (Basel). 2024;12(10):275. doi: 10.3390/sports12100275.

34. Wang B, Tan X, Yao X, Zhu Z. Sex-specific differences in the association of magne-sium intake with femoral neck bone mineral density among older adults. Endocr Connect. 2025;14(8):e250020. doi: 10.1530/EC-25-0020.

35. Mazza E, Maurotti S, Ferro Y, Castagna A, Pujia C, Sciacqua A, Pujia A, Montalcini T. Magnesium: Exploring Gender Differences in Its Health Impact and Dietary Intake. Nu-trients. 2025;17(13):2226. doi: 10.3390/nu17132226.

36. Obayashi M, Shimomura Y, Nakai N, Jeoung NH, Nagasaki M, Murakami T, Sato Y, Harris RA. Estrogen controls branched-chain amino acid catabolism in female rats. J Nutr. 2004;134(10):2628-33. doi: 10.1093/jn/134.10.2628.

37. Robbins R, O'Connor JC, Cortes TM, Serra MC. Branched-Chain Amino Acids Combined with Exercise Improves Physical Function and Quality of Life in Older Adults: Results from a Pilot Randomized Controlled Trial. Dietetics (Basel). 2025;4(3):32. doi: 10.3390/dietetics4030032.

38. Khemtong C, Kuo CH, Chen CY, Jaime SJ, Condello G. Does Branched-Chain Amino Acids (BCAAs) Supplementation Attenuate Muscle Damage Markers and Soreness after Resistance Exercise in Trained Males? A Meta-Analysis of Randomized Controlled Trials. Nutrients. 2021;13(6):1880. doi: 10.3390/nu13061880.

39. Bai GH, Tsai MC, Tsai HW, Chang CC, Hou WH. Effects of branched-chain amino acid-rich supplementation on EWGSOP2 criteria for sarcopenia in older adults: a systematic review and meta-analysis. Eur J Nutr. 2022;61(2):637-651. doi: 10.1007/s00394-021-02710-0.

40. D'Antona G, Ragni M, Cardile A, Tedesco L, Dossena M, Bruttini F, Caliaro F, Cor-setti G, Bottinelli R, Carruba MO, Valerio A, Nisoli E. Branched-chain amino acid supple-mentation promotes survival and supports cardiac and skeletal muscle mitochondrial bio-genesis in middle-aged mice. Cell Metab. 2010;12(4):362-372. doi: 10.1016/j.cmet.2010.08.016.

41. Zhang Y, Zhou Q, Yang R, Hu C, Huang Z, Zheng C, Liang Q, Gong R, Zhu X, Gong H, Yuan H, Chen C, Li X, Zhang N, Yang Z, Sun L. Serum branched-chain amino acids are associated with leukocyte telomere length and frailty based on residents from Guangxi longevity county. Sci Rep. 2020;10(1):10252. doi: 10.1038/s41598-020-67010-9.

42. van Dijk AM, Bruins Slot AS, Portincasa P, Siegerink SN, Chargi N, Verstraete CJR, de Bruijne J, Vleggaar FP, van Erpecum KJ. Systematic review with meta-analysis: Branched-chain amino acid supplementation in liver disease. Eur J Clin Invest. 2023;53(3):e13909. doi: 10.1111/eci.13909.

43. Noah L, Dye L, Bois De Fer B, Mazur A, Pickering G, Pouteau E. Effect of magne-sium and vitamin B6 supplementation on mental health and quality of life in stressed healthy adults: Post-hoc analysis of a randomised controlled trial. Stress Health. 2021;37(5):1000-1009. doi: 10.1002/smi.3051.

44. Moabedi M, Aliakbari M, Erfanian S, Milajerdi A. Magnesium supplementation bene-ficially affects depression in adults with depressive disorder: a systematic review and meta-analysis of randomized clinical trials. Front Psychiatry. 2023;14:1333261. doi: 10.3389/fpsyt.2023.1333261.

45. Khalid S, Mehboob R, Bokhari SS, Ali M, Shabbir A, Mehboob K, Adnan H, Karami MM, Shalabi H, Alshehri B. Comparative Efficacy of Magnesium and Potassium Towards Cholesterol and Quality of Life in Patients With Type 2 Diabetes Mellitus: A Randomised Single-Blinded Controlled Clinical Trial. Endocrinol Diabetes Metab. 2024;7(6):e511. doi: 10.1002/edm2.511.

46. Cepeda V, Ródenas-Munar M, García S, Bouzas C, Tur JA. Unlocking the Power of Magnesium: A Systematic Review and Meta-Analysis Regarding Its Role in Oxidative Stress and Inflammation. Antioxidants (Basel). 2025;14(6):740. doi: 10.3390/antiox14060740.

47. Han H, Fang X, Wei X, Liu Y, Jin Z, Chen Q, Fan Z, Aaseth J, Hiyoshi A, He J, Cao Y. Dose-response relationship between dietary magnesium intake, serum magnesium con-centration and risk of hypertension: a systematic review and meta-analysis of prospective cohort studies. Nutr J. 2017;16(1):26. doi: 10.1186/s12937-017-0247-4.

48. Rooney MR, Alonso A, Folsom AR, Michos ED, Rebholz CM, Misialek JR, Chen LY, Dudley S, Lutsey PL. Serum magnesium and the incidence of coronary artery disease over a median 27 years of follow-up in the Atherosclerosis Risk in Communities (ARIC) Study and a meta-analysis. Am J Clin Nutr. 2020;111(1):52-60. doi: 10.1093/ajcn/nqz256.

49. Qu X, Jin F, Hao Y, Li H, Tang T, Wang H, Yan W, Dai K. Magnesium and the risk of cardiovascular events: a meta-analysis of prospective cohort studies. PLoS One. 2013;8(3):e57720. doi: 10.1371/journal.pone.0057720.

50. Man JJ, Beckman JA, Jaffe IZ. Sex as a Biological Variable in Atherosclerosis. Circ Res. 2020;126(9):1297-1319. doi: 10.1161/CIRCRESAHA.120.315930.

51. Ravn HB, Korsholm TL, Falk E. Oral magnesium supplementation induces favorable antiatherogenic changes in ApoE-deficient mice. Arterioscler Thromb Vasc Biol. 2001;21(5):858-62. doi: 10.1161/01.atv.21.5.858.

52. Bundy JD, Cai X, Mehta RC, Scialla JJ, de Boer IH, Hsu CY, Go AS, Dobre MA, Chen J, Rao PS, Leonard MB, Lash JP, Block GA, Townsend RR, Feldman HI, Smith ER, Pasch A, Isakova T; CRIC Study Investigators. Serum Calcification Propensity and Clinical Events in CKD. Clin J Am Soc Nephrol. 2019 Nov 7;14(11):1562-1571. doi: 10.2215/CJN.04710419.

53. Mencke R, Al Ali L, de Koning MLY, Pasch A, Minnion M, Feelisch M, van Veld-huisen DJ, van der Horst ICC, Gansevoort RT, Bakker SJL, de Borst MH, van Goor H, van der Harst P, Lipsic E, Hillebrands JL. Serum Calcification Propensity Is Increased in Myo-cardial Infarction and Hints at a Pathophysiological Role Independent of Classical Cardio-vascular Risk Factors. Arterioscler Thromb Vasc Biol. 2024;44(8):1884-1894. doi: 10.1161/ATVBAHA.124.320974.

54. Eelderink C, Te Velde-Keyzer CA, Frenay AS, Vermeulen EA, Bachtler M, Aghag-olzadeh P, van Dijk PR, Gansevoort RT, Vervloet MG, Hillebrands JL, Bakker SJL, van Goor H, Pasch A, de Borst MH; NIGRAM2+ consortium. Serum Calcification Propensity and the Risk of Cardiovascular and All-Cause Mortality in the General Population: The PREVEND Study. Arterioscler Thromb Vasc Biol. 2020;40(8):1942-1951. doi: 10.1161/ATVBAHA.120.314187.

55. Bundy JD, Cai X, Mehta RC, Scialla JJ, de Boer IH, Hsu CY, Go AS, Dobre MA, Chen J, Rao PS, Leonard MB, Lash JP, Block GA, Townsend RR, Feldman HI, Smith ER, Pasch A, Isakova T; CRIC Study Investigators. Serum Calcification Propensity and Clinical Events in CKD. Clin J Am Soc Nephrol. 2019;14(11):1562-1571. doi: 10.2215/CJN.04710419.

56. Dahle DO, Åsberg A, Hartmann A, Holdaas H, Bachtler M, Jenssen TG, Dionisi M, Pasch A. Serum Calcification Propensity Is a Strong and Independent Determinant of Car-diac and All-Cause Mortality in Kidney Transplant Recipients. Am J Transplant. 2016;16(1):204-12. doi: 10.1111/ajt.13443.

57. Kurmann R, Buffle E, Pasch A, Seiler C, de Marchi SF. Predicting progression of aortic stenosis by measuring serum calcification propensity. Clin Cardiol. 2022;45(12):1297-1302. doi: 10.1002/clc.23922.

58. Bundy JD, Cai X, Scialla JJ, Dobre MA, Chen J, Hsu CY, Leonard MB, Go AS, Rao PS, Lash JP, Townsend RR, Feldman HI, de Boer IH, Block GA, Wolf M, Smith ER, Pasch A, Isakova T; CRIC Study Investigators. Serum Calcification Propensity and Coro-nary Artery Calcification Among Patients With CKD: The CRIC (Chronic Renal Insuffi-ciency Cohort) Study. Am J Kidney Dis. 2019;73(6):806-814. doi: 10.1053/j.ajkd.2019.01.024.

59. Kantauskaite M, Bolten K, Boschheidgen M, Schmidt C, Kolb T, Eckardt KU, Pasch A, Schimmöller L, Rump LC, Voelkl J, Stegbauer J. Serum Calcification Propensity and Calcification of the Abdominal Aorta in Patients With Primary Aldosteronism. Front Cardi-ovasc Med. 2022;9:771096. doi: 10.3389/fcvm.2022.771096.

60. Nollet L, Van Gils M, Fischer S, Campens L, Karthik S, Pasch A, De Zaeytijd J, Leroy BP, Devos D, De Backer T, Coucke PJ, Vanakker OM. Serum Calcification Propensi-ty T50 Associates with Disease Severity in Patients with Pseudoxanthoma Elasticum. J Clin Med. 2022;11(13):3727. doi: 10.3390/jcm11133727.

61.