Construction of UPLC£­Q£­TOF MS Fingerprints for Fingered Citron and Its Ingredient Differences Analysis before and after Processing

KeyWord:Fingered Citron  ultra-performance liquid chromatography£­quadrupole time-of-flight mass spectrometry£¨UPLC£­Q£­TOF MS£©  fingerprint  processing

Abstract:

      An ultra-performance liquid chromatography£­quadrupole time-of-flight mass spectrometry£¨UPLC£­Q£­TOF MS£© was applied to construct the fingerprints of raw Fingered Citron and processed Fingered Citron. Meanwhile£¬the chemical components differences between raw Fingered Citron and processed Fingered Citron were analyzed by chemical pattern recognition methods£®The sample was firstly ultrasonically extracted with methanol£¬and then separated on an Agilent Poroshell 120 EC£­C18£¨3.0 mm ¡Á 150 mm£¬2.7 ¦Ìm£© column by gradient elution£¬with 0.1% formic acid + 5 mmol/L ammonium acetate and acetonitrile as the mobile phases£®The MS system with electrospray ionization£¨ESI£© source was operated under positive and negative ion mode£®Based on the accurate mass£¬ fragment ions£¬ retention behaviors£¬ previous reports£¬net-work database and the fragmentation pathways£¬a total of 37 components in the raw Fingered Citron and the processed Fingered Citron were identified£¬including 10 flavonoids and flavonoid glycosides£¬4 coumarins£¬12 organic acids£¬5 glycosides£¬2 limonin and 4 other compounds£®The fingerprints for the two Fingered Citrons were established£¬and analyzed by a similar evaluation system for chromatographic fingerprint of traditional Chinese medicine£¨version 2004 A£¬National Committee of Pharmacopoeia£¬China£©£®The mass of each peak was accurately measured£¬and their chemical formula were generated via the MassHunter software£®The MS/MS spectrum for each peak was compared with the mass database and literature£¬and their chemical structures were deduced based on the MS/MS fragment£¬and finally verified by the reference substance£®In total£¬the UPLC£­Q£­TOF MS fingerprint for raw Fingered Citron contained 24 common peaks£¬of which 19 were identified. In addition£¬there were 22 common peaks in the UPLC£­Q£­TOF MS fingerprint for processed Fingered Citron£¬and 14 peaks were identified£®The principal component analysis£¨PCA£© and orthogonal partial£­least squares discriminant analysis£¨OPLS£­DA£© were adopted to analyze these data of two herbal medicines via SIMCA£­P software£¨pretreatment with Profinder software£©£®The results of PCA and OPLS£­DA showed that the two Fingered Citrons were significantly discriminated£®Nomilin acid£¬5-isopentenyloxy-7-methoxycoumarin£¬palmitic acid£¬oleic acid and oxypeucedanin hydrate were identified£¬and screened as the chemical markers that attributed to classification. Among them£¬the content of 5-isopentenyloxy-7-methoxycoumarin changed prominently£¬and was chosen as the chemical marker for differentiation on raw Fingered Citron and processed Fingered Citron£®This study provides a data support for the quality control of raw Fingered Citron and processed Fingered Citron.