Introduction

An attempt to speed up the simultaneous analysis of the 11 components of the UV absorber by using RHPLC resulted in the reduction of the analysis time to less than half that using conventional HPLC (refer to the Application Note No. 920013H).

LC-4000 RHPLC system

Experimental

HPLC Conditions
Column:   ZORBAX Eclipse Plus C8 (3.0 mm I.D. x 50 mm L, 1.8 µm)
Eluent A:   0.2 % formic acid
Eluent B:  Acetonitrile
Gradient:  A/B = 80/20 (0.00 min.) –> 35/65 (3.00 min.) –> 30/70 (6.50 min.) –>   5/95 (6.55 min.) –> 5/95 (7.00 min.) –>  80/20 (7.05 min.),1cycle; 9.5 min
Flow  rate:  0.8 mL/min
Column temp.:   40 ºC
Wavelength:   200 – 500 nm
Injection volume:   2 µL
Standard:  11 ultraviolet absorbers mixture in   acetonitrile (25 µg/mL each)
Sample:   Sunscreen cream
(Pretreatment method is described later.)

Keywords

UHPLC, PDA detector, ultraviolet absorbers, sunscreen cream, C8, EXTREMA

Results

Figure 1-1 shows the chromatogram and contour map of standard mixture of 11 ultraviolet absorbers obtained by a PDA detector. Figure 1-2 shows the spectra at each peak top.

Fig. 1-1   Chromatogram and contour map of ultraviolet absorbers standard mixture obtained by a PDA detector (25 µg/mL each).   (A): UV chromatogram (280, 310 nm), (B): contour map.
1: HMC, 2: HMBS, 3: THB, 4: EAB, 5: DHB, 6: DHDMB, 7: HMB, 8: BMB, 9: EMC, 10: EHS, 11: ECA.

Fig. 1-2   On-peak spectra of eleven ultraviolet absorbers.
1: HMC, 2: HMBS, 3: THB, 4: EAB, 5: DHB, 6: DHDMB, 7: HMB, 8: BMB, 9: EMC, 10: EHS, 11: ECA.

Figure 2 shows the procedure for extraction from sunscreen cream.

Fig. 2   Procedure for extraction from sunscreen cream

Figure 3 shows the chromatogram of extract from sunscreen cream. Figure 4 shows the spectral search results for sunscreen cream extract. The spectra of 11 components of standard sample were registered in the spectral library beforehand. By comparing between the spectrum of eluted peak in sunscreen cream extract and registered ones of standard sample, high correlation coefficient of 0.999 was obtained with EMC.

Fig. 3   Chromatogram of sunscreen
  (A) Standard, (B) Extraction.
1: HMC, 2: HMBS, 3: THB, 4: EAB, 5: DHB, 6: DHDMB, 7: HMB, 8: BMB, 9: EMC, 10: EHS, 11: ECA.

Fig. 4   Spectral search result for sunscreen cream extract
(A): Spectrum of Peak 9, (B): Spectrum of EMC.