VALIDASI METODE PENETAPAN KADAR ACID ORANGE 7 DAN SUDAN II DALAM SEDIAAN PERONA PIPI SECARA KCKT DAN SPEKTROSKOPI FTIR

NOVALINA BR PURBA, Prof. Dr. Abdul Rohman, M.Si., Apt. ; Prof. Dr. Sudibyo Martono M.S., Apt.

2019 | Tesis | MAGISTER ILMU FARMASI

Penggunaan Acid Orange 7 (AO7) sebagai pewarna dalam sediaan kosmetika diizinkan kecuali untuk pemakaian disekitar mata. Kemiripan warna dan ketersediaan dipasaran, menyebabkan kemungkinan dipalsukannya AO7 dengan Sudan II (SII). Penelitian tentang keamanan dan toksisitas Sudan II menyebabkan larangan penggunaannya pada kosmetika. Penjaminan mutu kosmetik membutuhkan metode yang valid, cepat murah dan sederhana seperti KCKT dan FTIR. Analisis Acid Orange 7 dan Sudan II dalam perona pipi dilakukan menggunakan kromatografi cair kinerja tinggi (KCKT) dan spektroskopi FTIR kombinasi kalibrasi multivariat partial least square (PLS). KCKT LC20AD (Shimadzu), kolom Thermo Hybrid Gold C18 (250x4,6 mm), 5 µm, detektor PDA pada panjang gelombang 482 nm (AO7) dan 495 nm (SII), fase gerak asetonitril:air menggunakan sistem gradien. Optimasi metode KCKT menggunakan metode optimasi Box-behnken Design (BBD), software Design Expert 7.1.5. Metode optimum divalidasi sesuai parameter ICH dalam hal selektivitas, linieritas, presisi, akurasi, batas deteksi dan batas kuantitasi. Analisis kuantitatif secara spektroskopi FTIR menggunakan teknik sampling ATR (Attenuated Total Reflectance), rentang bilangan gelombang scanning 4000-650 cm-1. Model kalibrasi diperoleh dengan memplot kadar aktual KCKT dengan kadar prediksi FTIR-PLS dari seri sampel lalu divalidasi internal dengan metode leave one out. Kriteria validasi berupa nilai root mean square error of calibration (RMSEC), root mean square error of cross validation (RMSECV), Root Mean Square Error of Predicted (RMSEP) yang semakin kecil dan R2 mendekati 1. Kondisi optimal KCKT hasil BBD adalah: konsentrasi ACN 1 43%, konsentrasi ACN 2 90%, laju alir 0,9 mL/menit dan suhu kolom 40oC. Hasil validasi metode analisis baik ditunjukkan oleh: metode selektif dengan nilai resolusi � 1,5, nilai R2 > 0.999 pada uji linearitas AO7 dan SII. Metode sensitif ditandai dengan nilai LOD dan LOQ rendah. Nilai RSD pada penetapan presisi intraday dan interday lebih rendah dari RSD Horwitz. Nilai akurasi AO7 dan SII pada level konsentrasi 80, 100 dan 120% berada di rentang 90-107%. Model kalibrasi FTIR yang diperoleh diukur pada bilangan gelombang 1500-550 cm-1. Model kalibrasi mempunyai nilai koefisien korelasi (R) baik yaitu 0,9999 (AO7) dan 0,9993 (SII). Hasil validasi internal leave one out RMSEC, RMSECV, RMSEP kecil dan R2 mendekati 1.

Acid Orange 7 (AO7) plays an importance role and allow as dye in cosmetic products except surround eyes use. Adulteration of AO7 with Sudan II (SII) because of color similarity and its availability in market. SII is banned for all cosmetics products based on safety and toxicity test. For ensuring cosmetic safety, it is needed valid, fast, inexpensive and easy method to analyse AO7 and SII such as HPLC and FTIR. Analysis of Acid Orange 7 and Sudan II in blusher was performed using high-performance liquid chromatography (HPLC) and FTIR spectroscopy combine by multivariate partial least square (PLS) calibration. HPLC analyse was performed by LC20AD (Shimadzu), Thermo Hybrid Gold C18 column (250x4,6 mm), 5 µm, PDA detector at wavelength 482 nm (AO7) dan 495 nm (SII), mobil phase acetonitrile:water was delivered by gradient system. Optimizing the HPLC method with Box-Behnken Design (BBD), Design Expert software 7.1.5. The optimum method was validated according to ICH parameters on selectivity, linearity, precision, accuracy, detection limit and limit of quantitation. Quantitative analysis using FTIR spectroscopy with ATR (Attenuated Total Reflectance) sampling technique. Calibration model performed by plotting the actual levels of HPLC with prediction levels of FTIR-PLS from the sample series. The model was validated internally. The validation criteria are root mean square error of calibration (RMSEC), root mean square error of cross validation (RMSECV), Root Mean Square Error of Predicted (RMSEP) and predicted residual error sum of square (PRESS) which are getting smaller and R2 approaching 1. Optimal condition of HPLC of BBD results are: ACN 1 concentration 43%, ACN 2 concentration 90%, flow rate of 0.9 mL/minute and column temperature 40oC. Results of good analysis method validation are shown by the selective method, R2 > 0.999 in the AO7 and SII linearity test. Sensitivy is characterized by low value of LOD and LOQ. RSD values on intraday and interday precision determination were lower than RSD Horwitz. Accuracy values of AO7 and SII at concentration levels of 80, 100 and 120% are in the range of 90-107%. Robustness lie in p value < 0,0001 (0,05). The calibration model obtained was measured at wave number 1500-550 cm-1. The calibration model has a good correlation coefficient (R) of 0.9999 (AO7) and 0.9993 (SII). Leave one out internal validation results, small RMSEC, RMSECV RMSEP and R2 close to 1.

Kata Kunci : Acid Orange 7, Sudan II, blusher, FTIR spectroscopy, multivariate PLS