• REYNA NEVITASARI Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta
  • Abdul Rohman Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta
  • SUDIBYO MARTONO Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Universitas Gadjah Mada, Yogyakarta



Carmine, Rhodamine B, RP-HPLC-PDA, Validation, Lipstick products


Objective: The objective of this study was to validate reversed phase-high performance liquid chromatography (RP-HPLC) method using photo-diode array detector for the simultaneous determination of carmine (CAR) and rhodamine B (RHO B) in lipstick products.

Methods: CAR and RHO B were analysed using Cosmosil C18 column (250 mm x 4.6 mm i.d., 5 µm) using Shimadzu LC 20AD chromatograph equipped with photo-diode array (PDA) detector at 245-600 nm. The mobile phase used was acetonitrile-phosphate buffer pH 3.4 (5.5:4.5 v/v) delivered isocratically at flow rate of 1.0 ml/min, using column temperature at 35 °C. The optimized HPLC condition was validated by determining several characteristics including selectivity, linearity, sensitivity expressed by the limit of detection and limit of quantification, precision and accuracy.

Results: The optimized HPLC condition could separate CAR and RHO B successfully with resolution value>2.0. The method was linear over studied concentration with a coefficient of determination (R2) 0.9999. The relative standard deviation (RSD) values were lower than those required RSD Horwitz and the recovery values were also acceptable, therefore, the developed method was precise and accurate.

Conclusion: HPLC with PDA detector has been accurately and successfully used for quantitative analysis of CAR and RHO B. The developed method is simple and can be used for routine analysis of CAR and RHO B for quality assurance purposes.


Download data is not yet available.


Nourmoradi H, Foroghi M, Farhadkhani M, Vahid Dastjerdi M. Assessment of lead and cadmium levels in frequently used cosmetic products in Iran. J Environ Public Health 2013;2013:962-727.

Wang J, Kay AB, Fletcher J, Formica MK, McAlindon TE. Is lipstick associated with the development of systemic lupus erythematosus (SLE)? Clin Rheumatol 2008;27:1183-7.

Kanekar H, Khale A. Coloring agents: current regulatory perspective for coloring agents intended for pharmaceutical and cosmetic use. Int J Pharm Phytopharmacol Res 2014;2:1–20.

Sabnis RW. Handbook of Biological Dyes and Stains: Synthesis and Industrial Applications, John Wiley and Sons, Inc., Hoboken, NJ, USA; 2010. p. 56.

Tatebe C, Zhong X, Ohtsuki T, Kubota H, Sato K, Akiyama H. A simple and rapid chromatographic method to determine unauthorized basic colorants (rhodamine B, auramine O, and pararosaniline) in processed foods. Food Sci Nutr 2014;2:547–56.

Greenhawt MJ, Baldwin JL. Carmine dye and cochineal extract: hidden allergens no more. Annals Allergy Asthma Immun 2009;103:73–5.

Fisher P. Review of using rhodamine b as a marker for wildlife studies. Wildlife Soc Bull 1999;27:318-29.

Rastogi SC, Barwick VJ, Carter SV. Identification of organic colourants in cosmetics by HPLC-diode array detection. Chromatographia 1997;45:215–28.

Miranda Bermudez E, Harp BP, Barrows JN. Qualitative Identification of permitted and non-permitted color additives in cosmetics. J AOAC Int 2014;97:1039–47.

Feng F, Zhao Y, Yong W, Sun L, Jiang G, Chu X. Highly sensitive and accurate screening of 40 dyes in soft drinks by liquid chromatography-electrospray tandem mass spectrometry. J Chromatogr B 2011;879:1813-8.

Liu X, Yu D, Yu Y, Ji S. Preparation of a magnetic molecularly imprinted polymer for selective recognition of rhodamine B. Appl Surface Sci 2014;320:138–45.

Gładysz M, Krol M, Koscielniak P. Differentiation of red lipsticks using the attenuated total reflection technique supported by two chemometric methods. Forensic Sci Int 2017;280:130–8.

Sharma S, Goyal S, Chauhan K. A review on analytical method development and validation. Int J Appl Pharm 2018;10:8-15.

Guidelines for validation of Analytical Procedures, Q2 (R1), ICH; 2005.

Kuber RB. Method development and validation for estimation of curcumin and piperine by RP-HPLC. Int J Appl Pharm 2019;11:216-21.

Cruz DD, Babu A, Joshy E, Anesh TP. Bioanalytical method development and validation of ticagrelor by RP-HPLC. Int J Appl Pharm 2017;9:51-4.

Miller JN, Miller JC. Statistics and chemometrics for analytical chemistry, Prentice Hall, England; 2010.

Gonzalez AG, Herrador MA. A practical guide to analytical method validation, including measurement uncertainty and accuracy profiles. Trends Anal Chem 2007;26:227-38.

Noviana E, Pranowo HD, Astuti A, Rohman A. Validation of mercury analyzer for determination of mercury in snake fruit. Int Food Res J 2012;18:933-6.



How to Cite

NEVITASARI, R., Rohman, A., & MARTONO, S. (2019). VALIDATION AND QUANTITATIVE ANALYSIS OF CARMINE AND RHODAMINE B IN LIPSTICK FORMULATION. International Journal of Applied Pharmaceutics, 11(3), 176–180.



Original Article(s)

Most read articles by the same author(s)

1 2 3 > >>