Traceability Of Wolfberries Based On Stable Isotope Ratio Mass Spectrometry

Isotope Ratio Mass Spectrometry（IRMS）technology,which can detect stable isotope information of substances,has been widely used in the fields of ecology,food,medicine,and justice in recent years.Stable isotopes can be used as fingerprint information for plant origin,but there are still many technical difficulties in the stable isotope analysis of specific compounds.The isotope fractionation effect is difficult to assess.Thus,a breakthrough in technology will have important implications for the identification of traditional Chinese medicines.Chinese wolfberry or goji berry（Lycium barbarum）is an important traditional Chinese medicine.Its function and price have a close correlation with its geographical provenance.Illegal mislabeling motivated by commercial gains brings serious food safety issues and damages consumer confidence.The literature reports on the use of chromatography,organic mass spectrometry,infrared spectroscopy,mineral element analysis and other techniques in the traceability of the origin of the wolfberry.Each analytical method has its advantages but also has some limitations.Therefore,it is very urgent and important to establish a new efficient,convenient,accurate and reliable traceability technology.In this work,a novel analytical strategy combined with chemometrics statistic tools was developed to determine the geographical origin of wolfberries from different provinces in China.Stable carbon isotopic ratios(δ¹³C_V-PDB)of wolfberry volatile compounds（i.e.limonene,tetramethylpyrazine,safranal,geranylacetone,andβ-ionone）were determined by gas chromatography-combustion-isotope ratio mass spectrometry（GC-C-IRMS）with headspace-solid phase micro extraction（HS-SPME）.They are all active or important sources of plant aroma.Five types of SPME fiber,extraction time,temperature and GC-IRMS conditions were comprehensively optimized to obtain the best adsorption of volatile compounds in wolfberry.Method integrity was assessed by comparing volatiles extracted using HS-SPME GC-IRMS with direct injection GC-IRMS and in good agreement（r=0.9997）with each other.The geographical variations of volatile compounds using one-way analysis of variance（ANOVA）were explored for individualδ¹³C_V-PDB-PDB values in wolfberry samples from Gansu,Ningxia and Qinghai.Geographical origin of wolfberry was differentiated by partial least squares-discrimination analysis（PLS-DA）,with an accuracy of 71.43%,85.71%and 100%for the three provinces,respectively.The results of linear discrimination analysis（LDA）is 89.16%,87.77%and 85.87%for these three provinces,respectively.In this work,a method was also established to measure the nitrogen isotope of amino acids by gas chromatography-combustion-isotope ratio mass spectrometry（GC-C-IRMS）.Twelve amino acids were derived with ethyl chloroformate as derivatized reagent.The precision of GC-C-IRMS is 0.13‰⁰.52‰,which fully meets the analytical accuracy requirements of isotopes.Besides,the results were highly correlated（r=0.990）with the element analysis-isotope ratio mass spectrometry（EA-IRMS）,indicating that no significant isotope fractionation was produced.The method can be used for the determination ofδ¹⁵N_Air values of amino acids in wolfberry.It can provide reference for further research on the traceability of wolfberry and other traditional Chinese medicines and the path of amino acid metabolism.Thus a fast,accurate and stable method for the stable isotope analysis of specific compounds in wolfberry was established in this research.Traceability studies of wolfberry can be performed by HS-SPME GC-C-IRMS combined with chemometric statistical analysis.Further,a method for determining the nitrogen stable isotope ratio of amino acid by GC-C-IRMS was established,which laid a foundation for more accurate traceability of traditional Chinese medicine origin.The research work of this thesis enriches the application of stable isotope analysis technology and provides a new technical means for the traceability of Chinese medicine origin.