Page 67 - Application Handbook - Liquid Chromatography
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Application  No.L491
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            n Quantitation of β-Carotene in Red Chili Pepper   separation obtained with the Nexera-e.
              by LC/MS/MS                                      Fig. 3 shows the two-dimensional separation data of
            The analytical conditions are shown in Table 1, and the   β-carotene obtained from DUIS-positive mode MRM
            sample pretreatment conditions are shown in Fig. 2.    analysis of the calibration curve, and Fig. 4 shows the
            β-carotene, which is a precursor of vitamin A, was   linearity of the three values (blobs) in the range of 0.01
            detected in the two-dimensional separation of the   to 10 mg/L, which correspond to the peak volumes
            carotenoids. Quantitation was then conducted using   used for quantitation. The correlation coefficient (r) =
            the LCMS-8030 triple quadrupole mass spectrometer.   0.998976 indicates results with good linearity. The
            Both high sensitivity and high selectivity can be   quantitative result for  β-carotene present in red chili
            obtained using MRM analysis, and further, reduced ion   pepper was calculated as 1.22 mg/L based on the
            suppression can be expected with the two-dimensional   concentration in the final sample following extraction.



                                                  Table 1  Analytical Conditions
                    1D Colum   : Ascentis Cyano (250 mm L. × 1.0 mm I.D., 5 µm)
                    Mobile Phase  : A; Hexane
                                 B; Hexane/Butylacetate/Acetone = 80/15/5 (v/v/v)
                    Flowrate   : 0.02 mL/min
                    Time Program  : B Conc. 0 % (0.01 min) → 0 % (5 min) → 100 % (65 min) → 100 % (75 min) → 0 % (76 min)
                    Column Temp. : 30 °C
                    Injection vol.  : 2 µL
                    Loop vol.  : 20 µL
                    2D Column  : Ascentis Express C18 (30 mm L. × 4.6 mm I.D., 2.7 µm)
                    Mobile Phase  : A; acetnitrile
                                 B; 2-propanol
                    Flowrate   : 4 mL/min (0.8 mL/min split for MS)
                    Time Program  : B Conc. 0 % (0.01 min) → 50 % (0.17 - 0.54 min) → 80 % (0.54 - 0.93 min) → 30 % (0.94 min) → STOP (1 min)
                    Detector   : SPD-M30A Photo diode array detector (standard cell, wave length = 450 nm)
                                                                                        536.40 > 444.30)
                                 Shimadzu LCMS-8030 (DUIS positive mode, targeted β-carotene MRM transition: m/z 536.40 > 444.30)
                                                                                     m/z
                                                                                     m/z



                     )PNPHFOBUFE SFE DIJMMJ QFQQFS
                        Capsicum annuum L.      H

               "EE DPOTFDVUJWF     N- BMJRVPUT PG NFUIBOPM
                 FUIZM BDFUBUF QFUSPMFVN FUIFS         W W W
                                                                                    Area = 551615
                      $PNCJOF FYUSBDUT BOE pMUSBUF                                 Concentration = 1.22 mg/L


                 &WBQPSBUF UP ESZOFTT VOEFS WBDVVN     …$

                                                                            Fig. 3  2D Plot of β-Carotene
                       %JTTPMWFE JO   N- NFUIBOPM
                      UFSU CVUZM NFUIZM FUIFS       W W


                     'JMUSBUF BOE JOKFDU   µ- JOUP )1-$

                                                                                         Correlation
                          Fig. 2  Sample Preparation                                     Coefficient (r) = 0.998976

                                                                                               Conc. (mg/L)

                                                                    Fig. 4  Linearity of Calibration Curve for β-Carotene




                                                        Data provided by University of Messina Prof. Luigi Mondello and Chromaleont S.r.l.

                                                                                                      First Edition: May. 2015


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