In vitro evaluation of the viability of vaginal cells (VK2/E6E7) and probiotic Lactobacillus species in lemon juice
Kingsley C. Anukam A B C and Gregor Reid AA Canadian Research and Development Centre for Probiotics, Lawson Health Research Institute, 268 Grosvenor Street, London, Ontario N6A4V2, Canada.
B Department of Pharmaceutical Microbiology, Faculty of Pharmacy, University of Benin, Nigeria.
C Corresponding author. Email: kanukam@uwo.ca
Sexual Health 6(1) 67-74 https://doi.org/10.1071/SH08037
Submitted: 21 May 2008 Accepted: 17 October 2008 Published: 23 February 2009
Abstract
Background: Women, especially in developing countries, most often bear the brunt of HIV infections. The continued lack of viable vaccines and microbicides has made some women resort to using natural products such as lemon or lime juice to avoid infection. Few in vitro studies have been done on the effect of lemon juice on vaginal cells and lactobacilli that constitute the major microbiota in healthy women. Objective: The objective of the present study was to evaluate in vitro the effect of lemon juice on the viability of vaginal cells (VK2/E6E7) and vaginal Lactobacillus species. Methods: Vaginal cells were exposed to different concentrations (0–30%) of lemon juice at pH 2.3 and 4.5 for 10 min. Viability was determined by staining the cells with propidium iodide and analysing them by flow cytometry. Lactobacillus organisms were dispensed into microplates with vaginally defined medium + peptone (VDMP) containing different concentrations of lemon juice ranging from 0 to 100%. Results: Lemon juice at pH 2.3 had a significant (P = 0.03) toxic effect on the vaginal cell line used. At 30% concentration, the vaginal cells were practically non-viable, typified by a 95% loss of viability, whereas at pH 4.5 there was only 5% cell loss. Lemon juice had varying growth inhibitory effects on the Lactobacillus species tested. At pH 4.5 and using 10–30% lemon juice, there was a stimulatory growth effect on certain Lactobacillus species. Conclusions: Lemon juice (20–30%) at pH 2.3 was highly toxic to VK2/E6E7 cells, and at pH 4.5 there was no significant effect on the viability of the cells within 10 min. Lemon juice above 10% at pH 2.3 was found to be detrimental to the growth of vaginal lactobacilli. Although lemon juice may be useful in other applications, its use in the vaginal region should be discouraged.
Additional keywords: bacteria, HIV, lemon juice, microbicides.
Acknowledgements
Dr Anukam is funded by the Ontario Neurotrauma Foundation and Dr Reid holds patent for Lactobacillus GR-1 and RC-14 as the most documented probiotic strains for women’s health.
[1] Imade GE, Sagay AS, Onwuliri VA, Egah DZ, Potts M, Short RV. Use of lemon or lime juice douches in women in Jos, Nigeria. Sex Health 2005; 2 237–9.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
[2] Imade G, Sagay A, Egah D, Onwuliri V, Grigg M, Egbodo C, et al. Prevalence of HIV and other sexually transmissible infections in relation to lemon or lime juice douching among female sex workers in Jos, Nigeria. Sex Health 2008; 5 55–60.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
[3] Stafford MK, Ward H, Flanagan A, Rosenstein IJ, Taylor-Robinson D, Smith JR, et al. Safety study of nonoxynol-9 as a vaginal microbicide: evidence of adverse effects. J Acquir Immune Defic Syndr Hum Retrovirol 1998; 17 327–31.
| PubMed |
[4] Thies FL, König W, König B. Rapid characterization of the normal and disturbed vaginal microbiota by application of 16S rRNA gene terminal RFLP fingerprinting. J Med Microbiol 2007; 56 755–61.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
[5] Hutchinson KB, Kip KE, Ness RB, Gynecologic Infection Follow-Through Investigators Vaginal douching and development of bacterial vaginosis among women with normal and abnormal vaginal microflora. Sex Transm Dis 2007; 34 671–5.
| Crossref | GoogleScholarGoogle Scholar |
[6] McClelland RS, Richardson BA, Graham SM, Masese LN, Gitau R, Lavreys L, et al. A prospective study of risk factors for bacterial vaginosis in HIV-1-seronegative African women. Sex Transm Dis 2008; 35 617–23.
[7] McClelland RS, Lavreys L, Hassan WM, Mandaliya K, Ndinya-Achola JO, Baeten JM. Vaginal washing and increased risk of HIV-1 acquisition among African women: a 10-year prospective study. AIDS 2006; 20 269–73.
[8] Fletcher PS, Harman SJ, Boothe AR, Doncel GF, Shattock RJ. Preclinical evaluation of lime juice as a topical microbicide candidate. Retrovirology 2008; 5 3.
| Crossref | GoogleScholarGoogle Scholar |
[9] Fichorova RN, Rheinwald JG, Anderson DJ. Generation of papillomavirus-immortalized cell lines from normal human ectocervical, endocervical, and vaginal epithelium that maintain expression of tissue-specific differentiation proteins. Biol Reprod 1997; 57 847–55.
| Crossref | GoogleScholarGoogle Scholar |
[10] Geshnizgani AM, Onderdonk AB. Defined medium simulating genital tract secretions for growth of vaginal microflora. J Clin Microbiol 1992; 30 1323–6.
[11] Hemmerling A, Potts M, Walsh J, Young-Holt B, Whaley K, Stefanski DA. Lime juice as a candidate microbicide? An open-label safety trial of 10% and 20% lime juice used vaginally. J Women’s Health 2007; 16 1041–51.
| Crossref | GoogleScholarGoogle Scholar |
[12] Annang L, Grimley DM, Hook EW 3rd. Vaginal douche practices among black women at risk: exploring douching prevalence, reasons for douching, and sexually transmitted disease infection. Sex Transm Dis 2006; 33 215–19.
| Crossref | GoogleScholarGoogle Scholar |
[13] Reid G, Jass J, Sebulsky MT, McCormick JK. Potential uses of probiotics in clinical practice. Clin Microbiol Rev 2003; 16 658–72.
| Crossref | GoogleScholarGoogle Scholar |
[14] Anukam KC, Koyama TE. Bile and acid tolerance of Lactobacillus plantarum KCA-1: a potential probiotical agent. Int J Dairy Sci 2007; 2 275–80.
[15] Myziuk L, Romanowski B, Johnson SC. BVBlue test for diagnosis of bacterial vaginosis. J Clin Microbiol 2003; 41 1925–8.
| Crossref | GoogleScholarGoogle Scholar |
[16] Nugent RP, Krohn MA, Hillier SL. Reliability of diagnosing bacterial vaginosis is improved by a standardized method of gram stain interpretation. J Clin Microbiol 1991; 29 297–301.
[17] Reid G, Beuerman D, Heinemann C, Bruce AW. Probiotic Lactobacillus dose required to restore and maintain a normal vaginal flora. FEMS Immunol Med Microbiol 2001; 32 37–41.
| Crossref | GoogleScholarGoogle Scholar |
[18] Boskey ER. Alternative therapies for bacterial vaginosis: a literature review and acceptability survey. Altern Ther Health Med 2005; 11 38–43.
[19] Drago L, De Vecchi E, Nicola L, Zucchetti E, Gismondo MR, Vicariotto F. Activity of a Lactobacillus acidophilus-based douche for the treatment of bacterial vaginosis. J Altern Complement Med 2007; 13 435–8.
| Crossref | GoogleScholarGoogle Scholar |
[20] Reid G, Beuerman D, Heinemann C, Bruce AW. Probiotic Lactobacillus dose required to restore and maintain a normal vaginal flora. FEMS Immunol Med Microbiol 2001; 32 37–41.
| Crossref | GoogleScholarGoogle Scholar |
[21] Anukam KC, Osazuwa E, Osemene GI, Ehigiagbe F, Bruce AW, Reid G. Clinical study comparing probiotic Lactobacillus GR-1 and RC-14 with metronidazole vaginal gel to treat symptomatic bacterial vaginosis. Microbes Infect 2006; 8 2772–6.
| Crossref | GoogleScholarGoogle Scholar |
