Synephrine in dietary supplements and specialized foodstuffs: biological activity, safety and methods of analysis

• Irina B. Perova
• Konstantin I. Eller
• Andrey V. Musatov
• Elena V. Tumolskaya

Abstract

Synephrine is a natural protoalkaloid of the bitter orange Citrus aurantium L., it has structural similarity to ephedrine and adrenaline. Synephrine in the form of bitter orange extract is widely used as an ingredient of dietary supplements (DS) and specialized foodstuffs (SF) intended for weight loss and fitness improvement. Along with thermogenic and lipolytic effects, synephrine can cause cardiovascular side effects, especially when combined with caffeine and physical activity. This aspect is important, insofar as the main consumers of weight loss products are overweight people who are at risk of developing cardiovascular diseases.

The aim of the research is a hygienic assessment of the usage of bitter orange extract and synephrine in DS and SF, which includes an analysis of approaches to technical regulation in the Russian Federation and abroad, a review of data on biological activity, safety, types of adulteration and methods for the determination of citrus protoalkaloids.

Results. The adrenergic effect of bitter orange is caused by the presence of R-(-)-p-synephrine, making up about 90% or more of the total protoalkaloids. Dry bitter orange fruit extracts, standardized to synephrine content, which can vary from 4 to 98%, are used in the production of DS and SF. Synephrine is a weak adrenergic agonist, acting primarily through β₃-adrenergic receptors, stimulating lipolysis. Because of insufficient safety data, the consumption of synephrine is regulated in the Russian Federation and abroad. The upper permissible level of synephrine consumption in the Russian Federation is 30 mg per day. Various cases of adulteration of DS and SF for weight loss and sport nutrition have been revealed: undeclared addition of synephrine in the form of bitter orange extract, addition of synthetic synephrine, its isomers or analogs. The main method for the determination of synephrine and other biogenic amines in DS and SF is high performance liquid chromatography with ultraviolet and/or mass detection.

Conclusion. The data presented in the review confirm the feasibility of developing an official method for determination of main protoalkaloids and monitoring of DS and SF for the content of synephrine and other citrus protoalkaloids on its basis.

Keywords:synephrine, bitter orange, dietary supplements, adulteration, HPLC

REFERENCES

1. Stohs S.J. Safety, efficacy, and mechanistic studies regarding Citrus aurantium (bitter orange) extract and p-synephrine. Phytother Res. 2017; 31 (10): 1463–74. DOI: https://doi.org/10.1002/ptr.5879

2. Ruiz-Moreno C., Del Coso J., Giráldez-Costas V., González-García J., Gutiérrez-Hellín J. Effects of p-synephrine during exercise: a brief narrative review. Nutrients. 2021; 13 (1): 233. DOI: https://doi.org/10.3390/nu13010233

3. Kaats G.R., Leckie R.B., Mrvichin N., Stohs S.J. Increased eating control and energy levels associated with consumption of bitter orange (p-synephrine) extract: a randomized placebo-controlled study. Nutr Diet Suppl. 2017; 9: 29–35. DOI: https://doi.org/10.2147/NDS.S136756

4. Di Lorenzo C., Ceschi A., Kupferschmidt H., Lüde S., De Souza Nascimento E., Dos Santos A., et al. Adverse effects of plant food supplements and botanical preparations: a systematic review with critical evaluation of causality. Br J Clin Pharmacol. 2015; 79( 4): 578–92. DOI: https://doi.org/10.1111/bcp.12519

5. EFSA Scientific Cooperation (ESCO) Working Group on Botanicals and Botanical Preparations. Advice on the EFSA guidance document for the safety assessment of botanicals and botanical preparations intended for use as food supplements, based on real case studies. EFSA J. 2009; 7 (9): 280. DOI: https://doi.org/10.2903/j.efsa.2009.280

6. EFSA SC (EFSA Scientific Committee), 2014. Scientific Opinion on a Qualified Presumption of Safety (QPS) approach for the safety assessment of botanicals and botanical preparations. EFSA J. 2014; 12 (3): 3593. DOI: https://doi.org/10.2903/j.efsa.2014.3593

7. Maisons-Alfort ANSES Opinion, Request No. 2012-SA-0200. OPINION of the French Agency for Food, Environmental and Occupational Health & Safetyon the risks associated with the presence in food supplements of p-synephrine or ingredients obtained from Citrus spp. fruits containing this substance. 2014: 46 p. URL: https://www.anses.fr/en/system/files/NUT2012sa0200EN.pdf

8. Stewart I., Newhall W.F., Edwads G.L. The isolation and identification of l-synephrine in leaves and fruit of Citrus. J Biol Chem. 1964; 239 (3): 930–2. DOI: https://doi.org/10.1016/s0021-9258(18)51679-3

9. Pellati F., Cannazza G., Benvenuti S. Study on the racemization of synephrine by off-column chiral high-performance liquid chromatography. J Chromatogr A. 2010; 1217 (21): 3503–10. DOI: https://doi.org/10.1016/j.chroma.2010.03.026.

10. Avula B., Upparapalli S.K., Navarrete A., Khan I.A. Simultaneous quantification of adrenergic amines and flavonoids in C. aurantium, various Citrus species, and dietary supplements by liquid chromatography. J AOAC Int. 2005; 88 (6): 1593–606.

11. BfR Opinion No. 004/2013. Health assessment of sports and weight loss products containing synephrine and caffeine. 2012: 29 p.

12. Food Standards Australia New Zealand (FSANZ). Risk assessment of oxedrine in foods intended to promote weight loss. 2015: 18 p.

13. Arbo M.D., Larentis E.R., Linck V.M., Aboy A.L., Pimentel A.L., Henriques A.T., et al. Concentrations of p-synephrine in fruits and leaves of Citrus species (Rutaceae) and the acute toxicity testing of Citrus aurantium extract and p-synephrine. Food Chem Toxicol. 2008; 46 (8): 2770–5. DOI: https://doi.org/10.1016/j.fct.2008.04.037

14. Stohs S.J., Preuss H.J. Stereochemical and pharmacological differences between naturally occuring p-synephrine and synthetic p-synephrine. J Funct Foods. 2012; 4 (1): 2–5. DOI: https://doi.org/10.1016/j.jff.2011.09.004

15. Sawler S. Health Canada Synephrine, Octopamine Caffeine Health Risk Assessment (HRA) Report. 2013: 49 p.

16. Tiesjema B., Jeurissen S.M.F., de Wit L., Mol H., Fragki S., Razenberg L. Risk assessment of synephrine. RIVM Report 2017-0069. 2017: 54 р. DOI: https://doi.org/10.21945/RIVM-2017-0069

17. Blumenthal M. Bitter orange peel and synephrine: Part 1 & Part 2. 2005: 28 p. URL: http://abc.herbalgram.org/site/DocServer/Bitter_Orange_Peel_and_Synephrine.pdf?docID=221

18. URL: https://rusada.ru/substances/prohibited-list/

19. ESCO report: Advice on the EFSA guidance document for the safety assessment of botanicals and botanical preparations intended for use as food supplements, based on real case studies. EFSA J. 2009; 7 (9): 280. DOI: https://doi.org/10.2903/j.efsa.2009.280; URL: https://www.ncaa.org/sport-science-institute/topics/2020-21-ncaa-banned-substances

20. URL: https://www.nutraingredients.com/Article/2018/03/22/Dutch-authorities-call-for-legal-limits-on-synephrine-in-food-supplements

21. URL: https://www.fda.gov/inspections-compliance-enforcement-and-criminal-investigations/compliance-actions-and-activities/warning-letters

22. Costa J.G., Vidovic B., Saraiva N., do Céu Costa M., Del Favero G., Marko D., et al. Contaminants: a dark side of food supplements? Free Radic Res. 2019; 53 (Suppl 1): 1113–35. DOI: https://doi.org/10.1080/10715762.2019.1636045

23. EU-28: Exporting Food Supplements to the European Union. 2017. GAIN Report No. E17004.

24. Pawar R.S., Sagi S., Leontyev D. Analysis of bitter orange dietary supplements for natural or synthetic phenethylamines by LC-MS/MS. Drug Test Anal. 2020; 12: 1241–51. DOI: https://doi.org/10.1002/dta.2871

25. Cohen P.A., Avula B., Venhuis B., Travis J.C., Wang Y.H., Khan I.A. Pharmaceutical doses of the banned stimulant oxilofrine found in dietary supplements sold in the USA. Drug Test Anal. 2017; 9 (1): 135–42. DOI: https://doi.org/10.1002/dta.1976

26. Roman M.C., Betz J.M., Hildreth J. Determination of synephrine in bitter orange raw materials, extracts, and dietary supplements by liquid chromatography with ultraviolet detection: single-laboratory validation. J AOAC Int. 2007; 90 (1): 68–81.

27. Lorenzo C.D., Santos A.D., Colombo F., Moro E., Dell’Agli M., Restani P. Development and validation of HPLC method to measure active amines in plant food supplements containing Citrus aurantium L. Food Control. 2014; 46: 136–42. DOI: https://doi.org/10.1016/J.FOODCONT.2014.05.017

28. Putzbach K., Rimmer C.A., Sharpless K.E., Sander L.C. Determination of Bitter Orange alkaloids in dietary supplements standard reference materials by liquid chromatography with ultraviolet absorbance and fluorescence detection. J Chromatogr A. 2007; 1156 (1–2): 304–11. DOI: https://doi.org/10.1016/j.chroma.2006.12.027.

29. Pharmacopoeia of the People’s Republic of China 2010 (English edition). 2010.

30. Taiwan Herbal Pharmacopoeia. 2th edition (English version). 2016: 44–5.

31. Sander L.C., Putzbach K., Nelson B.C., Rimmer C.A., Bedner M., Thomas J.B., et al. Certification of standard reference materials containing bitter orange. Anal Bioanal Chem. 2008; 391 (6): 2023–34. DOI: https://doi.org/10.1007/s00216-008-2074-0

32. Schaneberg B.T., Khan I.A. Quantitative and qualitative HPLC analysis of thermogenic weight loss products. Pharmazie. 2004; 59 (11): 819–23.

33. Ohta I., Mizunuma S., Yasuda T., Ohsawa K. Determination of synephrine in oriental pharmaceutical decoctions containing Evodiae fructus by ion-pair high-performance liquid chromatography. Yakugaku Zasshi. 1994; 114 (1): 33–8 (in Japanese). DOI: https://doi.org/10.1248/yakushi1947.114.1_33

34. Evans R.L., Siitonen P.H. Determination of caffeine and symphatomimetic alkaloids in weight loss supplements by high performance liquid chromatography. J Chromatogr Sci. 2008; 46 (1): 61–7. DOI: https://doi.org/10.1093/chromsci/46.1.61

35. Wagner H., Püls S., Barghouti T., Staudinger A., Melchart D. Chromatographic fingerprint analysis of herbal medicines. Volume 5. Thin-Layer and High Performance Liquid Chromatography of Chinese Drugs. Springer, 2018: 43–4. DOI: https://doi.org/10.1007/978-3-319-67062-1

36. Kim J.H., Kwak B.M., Ahn J.H., Park J.-S. Simultaneous determination of synephrine and N-methyltyramine in orange fruit and juice from Korean market by UPLC-FLD. Korean J Food Sci Technol. 2014; 46 (3): 276–82. DOI: http://dx.doi.org/10.9721/KJFST.2014.46.3.276

37. Pryce K., Samuel D., Lagares E., Myrthil M., Bess F., Harris A., et al. Presence of octopamine and an octopamine receptor in Crassostrea virginica. In vivo (Brooklyn). 2015; 37 (1): 16–24.

38. Li Q., Huang C., Huang Y. Sensitive determination of synephrine by flow-injection chemiluminescence. Luminescence. 2006; 21 (1): 43–8. DOI: https://doi.org/10.1002/bio.881

39. Liu Y., Xu F., Zhang Z., Yang C., Song R., Tian Y. Analysis of synephrine in da-cheng-qi decoction by HPLC employing precolumn derivatization with 9-fluorenylmethyl chloroformate. J Chromatogr Sci. 2009; 47 (10): 925–30. DOI: https://doi.org/10.1093/chromsci/47.10.925

40. Kobayashi Y., Yato M., Ito R., Saito K. Enantioselective determination of synephrine in health food products by liquid chromatography/time-of-flight mass spectrometry. Chromatography. 2020; 41: 39–44. DOI: https://doi.org/10.15583/jpchrom.2019.024

41. Tang Q., Zhang R., Zhou J., Zhao K., Lu Y., Zheng Y., et al. The levels of bioactive ingredients in Citrus aurantium L. at different harvest periods and antioxidant effects on H2O2-induced RIN-m5F cells. J Sci Food Agric. 2021; 101 (4): 1479–90. DOI: https://doi.org/10.1002/jsfa.10761

42. Bai Y., Zheng Y., Pang W., Peng W., Wu H., Yao H., et al. Identification and comparison of constituents of Aurantii fructus and Aurantii fructus immaturus by UFLC-DAD-triple TOF-MS/MS. Molecules. 2018; 23 (4): 803. DOI: https://doi.org/10.3390/molecules23040803

43. Shawky E. Determination of synephrine and octopamine in bitter orange peel by HPTLC with densitometry. J Chromatogr Sci. 2014; 52 (8): 899–904. DOI: https://doi.org/10.1093/chromsci/bmt113

44. Avula B., Upparapalli S.K., Khan I.A. Enantiomeric separation of adrenergic amines in Citrus species, related genera and dietary supplements by capillary electrophoresis. Chromatographia. 2005; 62 (3): 151–7. DOI: https://doi.org/10.1365/s10337-005-0598-4

45. Yi Y.N., Cheng X.M., Liu L.A., Hu G.Y., Wang Z.T., Deng Y.D., et al. Simultaneous determination of synephrine, arecoline, and norisoboldine in Chinese patent medicine Si-Mo-Tang oral liquid preparation by strong cation exchange high performance liquid chromatography. Pharm Biol. 2012; 50 (7): 832–8 DOI: https://doi.org/10.3109/13880209.2011.637505

46. Zhang Y., Jiao B. Simultaneous determination of six protoalkaloids in Chinese local varieties of loose-skin mandarins and sweet oranges by strong cation exchange-high performance liquid chromatography. Food Anal Methods. 2019; 12: 677–86. DOI: https://doi.org/10.1007/s12161-018-1401-0

47. URL: https://www.sielc.com/Application-HPLC-Separation-of-Octopamine-and-Synephrine-on-Primesep-200-column.html

48. Tanaka S., Sekiguchi M., Yamamoto A., Aizawa S.I., Sato K., Taga A., et al. Separation of synephrine enantiomers in citrus fruits by a reversed phase HPLC after chiral precolumn derivatization. Anal Sci. 2019; 35 (4): 407–12. DOI: https://doi.org/10.2116/analsci.18P441

All articles in our journal are distributed under the Creative Commons Attribution 4.0 International License (CC BY 4.0 license)