Serum proteins differentially expressed in gestational diabetes mellitus assessed using isobaric tag for relative and absolute quantitation proteomics

BACKGROUND As a well-known fact to the public,gestational diabetes mellitus(GDM)could bring serious risks for both pregnant women and infants.During this important investigation into the linkage between GDM patients and their altered expression in the serum,proteomics techniques were deployed to detect the differentially expressed proteins(DEPs)of in the serum of GDM patients to further explore its pathogenesis,and find out possible biomarkers to forecast GDM occurrence.METHODS Subjects were divided into GDM and normal control groups according to the IADPSG diagnostic criteria.Serum samples were randomly selected from four cases in each group at 24-28 wk of gestation,and the blood samples were identified by applying iTRAQ technology combined with liquid chromatography-tandem mass spectrometry.Key proteins and signaling pathways associated with GDM were identified by bioinformatics analysis,and the expression of key proteins in serum from 12 wk to 16 wk of gestation was further verified using enzyme-linked immunosorbent assay (ELISA).RESULTS Forty-seven proteins were significantly differentially expressed by analyzing the serum samples between the GDMgravidas as well as the healthy ones. Among them, 31 proteins were found to be upregulated notably and the rest16 proteins were downregulated remarkably. Bioinformatic data report revealed abnormal expression of proteinsassociated with lipid metabolism, coagulation cascade activation, complement system and inflammatory responsein the GDM group. ELISA results showed that the contents of RBP4, as well as ANGPTL8, increased in the serumof GDM gravidas compared with the healthy ones, and this change was found to initiate from 12 wk to 16 wk ofgestation.CONCLUSION GDM symptoms may involve abnormalities in lipid metabolism, coagulation cascade activation, complementsystem and inflammatory response. RBP4 and ANGPTL8 are expected to be early predictors of GDM.

LC-MS/MS法定量测定人血浆中阿司匹林及其代谢产物水杨酸

A sensitive liquid chromatography-electrospray ionization-tandem mass spectrometric (LC-MS/MS) method for the simultaneous determination of aspirin (ASA) and its metabolite salicylic acid (SA) in human plasma has been developed and validated.The plasma ASA and SA were extracted using a liquid-liquid extraction (LLE) and the sample extract was injected onto the LC-MS/MS system.The limits of quantitation(LLOQ) for ASA and SA are both 25 ng/mL.This method allowed the reproducible and accurate quantification with the concentration rang of 25-10 000 ng/mL.This method could be applied to the quantitation aspirin and salicylic acid in human plasma.

液相色谱同位素稀释质谱对寡核苷酸含量的测定研究

The mass of oligonucleotides was quantitated by liquid chromatography-isotope dilution mass spectrometry.The uncertainty was calculated.This could provide information for the further quantitative measurement and utilization of DNA.

液相色谱-同位素稀释质谱精确测定奶粉中的核苷酸含量

Adenosine 5′-monophosphate in powdered milk was quantitated by liquid chromatography-isotope dilution mass spectrometry.The uncertainty was calculated.This could provide information for the further quantitative measurement and utilization of 5′-nucleotide acid as food additive.

Identification and quantification of the bioactive components in Osmanthus fragrans roots by HPLC-MS/MS

The roots of O. fragrans are also a valuable resource in addition to its flowers and fruits. In this study, the HPLC-MS/MS method used for analyzing the chemical constituents in O. fragrans roots extract was developed, which showed high sensitivity for both qualitative and quantitative analyses. Thirty-two compounds were first discovered in O. fragrans roots, one compound of which was reported for the first time. The simultaneous determination method for acteoside, isoacteoside, oleuropein and phillyrin was validated to be sensitive and accurate. Then it was applied to determine the content of bioactive components in O. fragrans roots from different cultivars. The content of oleuropein and phillyrin in the twelve batches was relatively stable, while the content of acteoside and isoacteoside varied greatly.Moreover, the therapeutic material basis and mechanism of O. fragrans roots exerting its traditional pharmacodynamics were analyzed by network pharmacology. The results showed that O. fragrans roots might be effective for the treatment of inflammation, cardiovascular diseases, cancer, and rheumatoid arthritis, which is consistent with the traditional pharmacodynamics of O. fragrans roots. This work can provide an analytical method for the comprehensive development of O. fragrans roots.

The ＇omics revolution and our understanding of sperm cell biology

The foundations of proteomics are to study gene products and their regulatory roles within cells. Paradoxically, the only evidence that sperm cells make new proteins is through mitochondrial protein synthesis. Yet despite this, spermatozoa are the perfect candidates for mass spectrometry and hence, proteomic analysis. These enterprising cells use a plethora of post-translational modifications in order to gain functionality following their production within the testis. By using a combination of two-dimensional polyacrylamide gel electrophoresis （2D-PAGE）, and more recently liquid chromatography-mass spectrometry （LC-MS）/MS, recent advances in sperm cell biology, through the use of proteomics, is making unparalleled progress. The protein inventory lists being generated have shed light on transmembrane proteins, kinases and chaperones never previously recognized. In addition, the ability to isolate either phosphopeptides or glycopeptides and quantify the differences between cells of two different populations make proteomic analysis of spermatozoa a real chance to finally answer some age old questions.

Quantitation of HBsAg predicts response to entecavir therapy in HBV genotype C patients

AIM： To analysis the factors that predict the response to entecavir therapy in chronic hepatitis patients with hepatitis B virus （HBV） genotype C. METHODS： Fifty patients [hepatitis B e antigen （HBeAg）- negative：HBeAg-positive = 26：24] with HBV genotype C, who received nalve entecavir therapy for 〉 2 years, were analyzed. Patients who showed HBV DNA levels ≥ 3.0 log viral copies/mL after 2 years of entecavir ther- apy were designated as slow-responders, while those that showed 〈 3.0 log copies/mL were termed rapid- responders. Quantitative hepatitis B surface antigen （HBsAg） levels （qHBsAg） were determined by the Archi- tect HBsAg QT immunoassay. Hepatitis B core-related antigen was detected by enzyme immunoassay. Pre-C and Core promoter mutations were determined using by polymerase chain reaction （PCR）. Drug-resistance muta- tions were detected by the PCR-Invader method. RESULTS： At year 2, HBV DNA levels in all patients in the HBeAg-negative group were 〈 3.0 log copies/mL. In contrast, in the HBeAg-positive group, 41.7% were slow-responders, while 58.3% were rapid-responders. No entecavir-resistant mutants were detected in the slow-responders. When the pretreatment factors were compared between the slow- and rapid-responders; the median qHBsAg in the slow-responders was 4.57 log IU/mL, compared with 3.63 log IU/mL in the rapid- responders （P 〈 0.01）. When the pretreatment factors predictive of HBV DNA-negative status at year 2 in all 50 patients were analyzed, HBeAg-negative status, low HBV DNA levels, and low qHBsAg levels were signifi- cant （P 〈 0.01）. Multivariate analysis revealed that the low qHBsAg level was the most significant predictive factor （P = 0.03）. CONCLUSION： Quantitation of HBsAg could be a use- ful indicator to predict response to entecavir therapy.

Identification of protein targets for the antidepressant effects of Kai-Xin-San in Chinese medicine using isobaric tags for relative and absolute quantitation

Kai-Xin-San consists of Ginseng Radix, Polygalae Radix, Acori Tatarinowii Rhizoma, and Poria at a ratio of 3:3:2:2. Kai-Xin-San has been widely used for the treatment of emotional disorders in China. However, no studies have identified the key proteins implicated in response to Kai-Xin-San treatment. In this study, rat models of chronic mild stress were established using different stress methods over 28 days. After 14 days of stress stimulation, rats received daily intragastric administrations of 600 mg/kg Kai-Xin-San. The sucrose preference test was used to determine depression-like behavior in rats, while isobaric tags were used for relative and absolute quantitation-based proteomics to identify altered proteins following Kai-Xin-San treatment. Kai-Xin-San treatment for 2 weeks noticeably improved depression-like behaviors in rats with chronic mild stress. We identified 33 differentially expressed proteins: 7 were upregulated and 26 were downregulated. Functional analysis showed that these differentially expressed proteins participate in synaptic plasticity, neurodevelopment, and neurogenesis. Our results indicate that Kai-Xin-San has an important role in regulating the key node proteins in the synaptic signaling network, and are helpful to better understand the mechanism of the antidepressive effects of Kai-Xin-San and to provide objective theoretical support for its clinical application. The study was approved by the Ethics Committee for Animal Research from the Chinese PLA General Hospital(approval No. X5-2016-07) on March 5, 2016.

Different protein expression patterns in rat spinal nerves during wallerian degeneration assessed using isobaric tags for relative and absolute quantitation proteomics profiling

Sensory and motor nerve fibers of peripheral nerves have different anatomies and regeneration functions after injury. To gain a clear understanding of the biological processes behind these differences, we used a labeling technique termed isobaric tags for relative and absolute quantitation to investigate the protein profiles of spinal nerve tissues from Sprague-Dawley rats. In response to Wallerian degeneration, a total of 626 proteins were screened in sensory nerves, of which 368 were upregulated and 258 were downregulated. In addition, 637 proteins were screened in motor nerves, of which 372 were upregulated and 265 were downregulated. All identified proteins were analyzed using the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis of bioinformatics, and the presence of several key proteins closely related to Wallerian degeneration were tested and verified using quantitative real-time polymerase chain reaction analyses. The differentially expressed proteins only identified in the sensory nerves were mainly relevant to various biological processes that included cell-cell adhesion, carbohydrate metabolic processes and cell adhesion, whereas differentially expressed proteins only identified in the motor nerves were mainly relevant to biological processes associated with the glycolytic process, cell redox homeostasis, and protein folding. In the aspect of the cellular component, the differentially expressed proteins in the sensory and motor nerves were commonly related to extracellular exosomes, the myelin sheath, and focal adhesion. According to the Kyoto Encyclopedia of Genes and Genomes, the differentially expressed proteins identified are primarily related to various types of metabolic pathways. In conclusion, the present study screened differentially expressed proteins to reveal more about the differences and similarities between sensory and motor nerves during Wallerian degeneration. The present findings could provide a reference point for a future investigation into the differences between sensory and motor nerves in Wallerian degeneration and the characteristics of peripheral nerve regeneration. The study was approved by the Ethics Committee of the Chinese PLA General Hospital, China(approval No. 2016-x9-07) in September 2016.

A 3-D morphological approach on spatial form and cultural identity of ethnic mountain settlements:Case from Guizhou,China

Ethnic mountain settlements are living heritage of varied vernacular cultures.The preservation of both the built form and the intangible socio-cultural associations with them are global concern in process of urbanization,and in the notion of sustainable development.However,there is a lack of multi-dimensional and cross-cultural quantitative research in settlement morphology,making it difficult to guide practice effectively.Therefore,this study focuses on exploring an automatic or semi-automatic quantification and classification method for the morphological identity of ethnic mountain settlements.We introduce and combine 3-D morphological indicators with existing 2-D indicators to build and test three different sets of indication systems for semi-automatic classification for the settlements’ethnic attribute basing on spatial morphology.Taking the Miao,Dong,and Tunpu(Han)ethnic settlements in Guizhou province,southwest China as research samples,we applied factor analysis and hierarchical clustering methods to compare the classification accuracy under the three systems using data from topographic map,field investigation map,satellite imageries,and ethnography or local chronicle.The results showed that,the 3-D indication system has succeeded in semi-automatic quantification and classification of settlement morphology and ethnic identity by greatly increasing the classification accuracy to 96.30%,which is a huge improvement compared with the basic 2-D indication system(42.59%)and the advanced 2-D indication system(61.11%).The settlement samples are further divided into two sub-types with significant morphological differences in each major ethnic category under the 3-D indication system.We then discussed the potential improvement and future large-scale application of this method with the help of machine learning and other smart techniques.We hope to provide a comprehensive quantitative perspective and a more scientific reference for the future preservation and sustainable development of the massive and diverse vernacular heritages across the world.

Improved image resolution on thoracic carcinomas by quantitative 18F-FDG coincidence SPECT/CT in comparison to 18F-FDG PET/CT

Currently,18F-FDG coincidence SPECT(Co-SPECT)/CT scan still serves as an important tool for diagnosis,staging,and evaluation of cancer treatment in developing countries.We implemented full physical corrections(FPC) to Co-SPECT(quantitative Co-SPECT) to improve the image resolution and contrast along with the capability for image quantitation.FPC included attenuation,scatter,resolution recovery,and noise reduction.A standard NEMA phantom filled with 10:1 F-18 activity concentration ratio in spheres and background was utilized to evaluate image performance.Subsequently,15 patients with histologically confirmed thoracic carcinomas were included to undergo a 18 F-FDG Co-SPECT/CT scan followed by a 18 F-FDG PET/CT scan.Functional parameters as SUVmax,SUVmean,SULpeak,and MTV from both quantitative Co-SPECT and PET were analyzed.Image resolution of Co-SPECT for NEMA phantom was improved to reveal the smallest sphere from a diameter of 28 mm to 22 mm(17 mm for PET).The image contrast was enhanced from 1.7 to 6.32(6.69 for PET) with slightly degraded uniformity in background(3.1% vs.6.7%)(5.6% for PET).Patients’ SUVmax,SUVmean,SULpeak,and MTV measured from quantitative Co-SPECT were overall highly correlated with those from PET(r=0.82-0.88).Adjustment of the threshold of SUVmax and SUV to determine SUVmean and MTV did not further change the correlations with PET(r=0.81-0.88).Adding full physical corrections to Co-SPECT images can significantly improve image resolution and contrast to reveal smaller tumor lesions along with the capability to quantify functional parameters like PET/CT.

Identification and Quantitation of Cashmere (Pashmina) Fiber and Wool Using Novel Microchip Based Real-Time PCR Technology

The textile industrial chain all over the world is facing a challenge of differentiating cashmere fiber from mixture of wool and other fibers in case cashmere stocks are adulterated with wool or other fibers. For identification of cashmere in such mixtures, the development of microchip based real-time PCR technology offers a very sensitive, specific, and accurate solution. The technology has been validated with cashmere and wool samples procured from distant farms, and from cashmere goats and sheep of different age and sex. Model samples with incremental raw cashmere or wool content were tested. The experimentally determined content was found to be comparable to the weighed content of the respective fibers in the samples. This technology may prove a cost cutter since it needs only 1.2 μl of the PCR reagent mix. It is substantially faster than traditional real-time PCR systems for being carried as miniature reaction volume in metal microchip. These features allow faster thermal equilibrium and thermal uniformity over the entire array of microreactors. For routine tests or in commercial set up, the microchips are available as ready-to-run with lyophilized reagents in its microreactors to which only 1 μl of the 10-fold diluted isolated DNA sample is added. The lyophilized microchips offer user-friendly handling in testing laboratories and help minimize human error.

Non-invasive determination of hepatic steatosis by acoustic structure quantification from ultrasound echo amplitude

AIM:To use leptin-deficient(ob/ob) mice with demonstrated differences in steatosis levels to test a new diagnostic method using the acoustical structure quantification(ASQ) mode and the associated analytical parameter,"focal disturbance ratio"(FD-ratio).METHODS:Nine ob/ob mice,at 5,8,and 12 wk of age(n = 3 in each age group),were used as models for hepatic steatosis.Echo signals obtained from ultrasonography in the mice were analyzed by ASQ,which uses a statistical analysis of echo amplitude to estimate inhomogeneity in the diagnostic region.FD-ratio,as calculated from this analysis,was the focus of the present study.FD-ratio and fat droplet areas and sizes were compared between age groups.RESULTS:No fibrosis or inflammation was observed in any of the groups.The fat droplet area significantly(P < 0.01) increased with age from 1.25% ± 0.28% at 5 wk to 31.07% ± 0.48% at 8 wk to 51.69% ± 3.19% at 12 wk.The median fat droplet size also significantly(P < 0.01) increased with age,from 1.33(0.55-10.52) m at 5 wk,2.82(0.61-44.13) m at 8 wk and 6.34(0.66-81.83) m at 12 wk.The mean FD-ratio was 0.42 ± 0.11 at 5 wk,0.11 ± 0.05 at 8 wk,and 0.03 ± 0.02 at 12 wk.The FD-ratio was significantly lower at 12 wk than at 5 wk and 8 wk(P < 0.01).A significant negative correlation was observed between the FD-ratio and either the fat droplet area(r =-0.7211,P = 0.0017) or fat droplet size(r =-0.9811,P = 0.0052).CONCLUSION:This tool for statistical analysis of signals from ultrasonography using the FD-ratio can be used to accurately quantify fat in vivo in an animal model of hepatic steatosis,and may serve as a quantitative biomarker of hepatic steatosis.

Comparison of Abbott and Da-an real-time PCR for quantitating serum HBV DNA

AIM: To compare the performance of the Da-an real-time hepatitis B virus (HBV) DNA assay and Abbott RealTime HBV assay.

Quantitation of Genital Herpes Virus DNA by Polymerase Chain Reaction and ELISA

Objective: To detect and quantitate genital herpes simplexvirus (HSV) DNA in specimens from 100 patients clinicallydiagnosed with genital herpes. Methods: Polymerase Chain Reaction (PCR) andenzyme-linked immunosorbent assay (ELISA) were used witha standard curve of DNA copies of HSV as quantitativecontrast. Results: Ninety-three cases were confirmed HSV positiveand 7 cases were found to be negative. There were 58 cases ofHSV-2 (62.4%) and 35 cases of HSV-1(37.6%) among the 93positive cases. The number of DNA plasmids ranged from 115to 1.1×10~5 per 250μL among the 93 positive samples (mean=7.1×10~4/250μL). The number of HSV DNA plasmids rangedfrom 136 to 1.1×10~5 copies per 250μL(mean=7.6×10~4) amongthose with HSV-2, and 115 to 9.4×10~4 per 250μL(mean=6.3×10~4) among those with HSV-1. Meanwhile 10μL ofextracted and dissolved DNA randomly taken from 8 each ofHSV-2 and HSV-1 samples were tested. The number of HSV-2DNA plasmids ranged from 35 copies to 2.7×10~4 (Mean=1.8×10~4) and the number of HSV-1 DNA ranged from 29 to2.5×10~4 (Mean=1.6×10~4). In the 7 negative cases, the quantityof HSV plasmids was zero. Conclusion: The sensitivity of ELISA quantitation (93%) isequal to that of Southern blot. The sensitivity of PCR fordiagnosis is 91%, and 88% for PCR typing.

Non-target Screening and Quantitation of Organic Chlorides in Oilfield Chemicals with Comprehensive Two Dimensional Gas Chromatography Coupled with Time of Flight Mass Spectrometry

Advanced analytical methodology based on comprehensive two-dimensional gas chromatography-time of flight mass spectrometry(GC×GC-TOFMS) is proposed for investigation of organic chlorides in different oilfield chemicals.The target screening was initially carried out on 8 suspected organic chlorides by evaluating the capability of the enhanced separation and reliable identification at a trace concentration. GC×GC-TOFMS allowed for the fast and automated analysis of organic chlorides at a level of 200 μg/L. This method was subsequently applied for non-target screening of organic chlorides in different oilfield chemicals at various locations across China. 22 organic chlorides were identified and verified by comparison with pure standards in the mixed sample. Finally, this method was used to determine the content of the organic chlorides in individual samples. The result showed that the organic chloride levels in 19 of the 39 tested oilfield chemicals were above the threshold limit of 1.0 mg/L.

Simultaneous Determination of Catechins and Caffeine in Green Tea-Based Beverages and Foods for Specified Health Uses

Catechins in green tea have various useful features including antioxidant activity and preventive effects on metabolic syndrome. Various beverages that are enriched with tea catechins are marketed as Foods for Specified Health Uses (FOSHU) in Japan. However, recent reports have indicated that excessive consumption of green tea extracts as a dietary supplement are associated with adverse health effects such as liver disorders. Various catechins and caffeine are constituents of FOSHU tea-based beverages. The amount of catechins in FOSHU products is displayed on labels as total catechin content, but the content of individual catechins are not provided. Although health hazards of FOSHU products have rarely been reported, precise information about the content and types of catechins in FOSHU products is needed to ensure safety. We used high-performance liquid chromatography with a photodiode array (HPLC/PDA) to simultaneously identify and quantify catechins and caffeine in green tea-based popular beverages and FOSHU beverages. This technique allowed simultaneous quantitation of five types of catechins and caffeine in green tea without complicated sample preparation. Epigallocatechin gallate (EGCG) and epigallocatechin EGC were the main catechins in various FOSHU beverages and the concentrations of almost all catechins were higher in FOSHU, than in popular green tea-based beverages. The concentrations of EGCG in green tea-based popular beverages and in FOSHU beverages were 5.4 - 7.3 and 10.2 - 41.9 mg/100mL, respectively, with the highest concentration being in a product named Healthya (approximately 147 mg/bottle). The simultaneous determination of compounds such as catechins and caffeine in FOSHU beverages can help to estimate beneficial and adverse effects to prevent deleterious effects on health and the excessive consumption of FOSHU beverages containing high concentrations of tea catechins should be avoided.

Simultaneous determination of kolliphor HS15 and miglyol 812 in microemulsion formulation by ultra-high performance liquid chromatography coupled with nano quantity analyte detector

A novel method for simultaneous determination of kolliphor HS15 and miglyol 812 in microemulsion formulation was developed using ultra-high performance liquid chromatography coupled with a nano quantitation analytical detector （UHPLC-NQAD）. All components in kolliphor HS15 and miglyo1812 were well separated on an Acquity BEH C18 column. Mobile phase A was 0.1% trifluoroacetic acid （TFA） in water and mobile phase B was acetonitrile. A gradient elution sequence was programed initially with 60% organic solvent, slowly increased to 100% within 8 min. The flow rate was 0.7 mL/min. Good linearity （r 〉 0.95） was obtained in the range of 27.6-1381.1 μg/mL for polyoxyl 15 hydroxystearate in kolliphor HS15, 0.8-202.0 μg/mL for caprylic acid triglyceride and 2.7-221.9μg/mL for capric acid triglyceride in miglyol 812. The relative standard deviations （RSD） ranged from 0.6% to 1.7% for intra-day precision and from 0.4% to 2.7% for inter-day precision. The overall recoveries （accuracy） were 99.7%-101.4% for polyoxyl 15 hydroxystearate in kolliphor HS15, 96.7%-99.6% for caprylic acid triglyceride, and 94.1%- 103.3% for capric acid triglyceride in miglyol 812. Quantification limits （QL） were determined as 27.6 μg/ mL for polyoxy115 hydroxystearate in kolliphor HS15, 0.8 μg/mL for caprylic acid triglyceride, and 2.7 μg/ mL for capric acid triglyceride in miglyol 812. No interferences were observed in the retention time ranges of kolliphor HSI5 and miglyol 812. The method was validated in terms of specificity, linearity, precision, accuracy, QL, and robustness. The proposed method has been applied to microemulsion for- mulation analyses with good recoveries （82.2%-103.4%）.

Proteomic analysis of trans-hemispheric motor cortex reorganization following contralateral C7 nerve transfer

Nerve transfer is the most common treatment for total brachial plexus avulsion injury. After nerve transfer, the movement of the injured limb may be activated by certain movements of the healthy limb at the early stage of recovery, i.e., trans-hemispheric reorganization. Pre- vious studies have focused on functional magnetic resonance imaging and changes in brain-derived neurotrophic factor and growth asso- ciated protein 43, but there have been no proteomics studies. In this study, we designed a rat model of total brachial plexus avulsion injury involving contralateral C7 nerve transfer. Isobaric tags for relative and absolute quantitation and western blot assay were then used to screen differentially expressed proteins in bilateral motor cortices. We found that most differentially expressed proteins in both cortices of upper limb were associated with nervous system development and function （including neuron differentiation and development, axonogenesis, and guidance）, microtubule and cytoskeleton organization, synapse plasticity, and transmission of nerve impulses. Two key differentially expressed proteins, neurofilament light （NFL） and Thy-1, were identified. In contralateral cortex, the NFL level was upregulated 2 weeks after transfer and downregulated at 1 and 5 months. The Thy-1 level was upregulated from 1 to 5 months. In the affected cortex, the NFL level increased gradually from 1 to 5 months. Western blot results of key differentially expressed proteins were consistent with the proteom- ic findings. These results indicate that NFL and Thy-1 play an important role in trans-hemispheric organization following total brachial plexus root avulsion and contralateral C7 nerve transfer.

Rapid Quantitative Determination of Isoprene Monomer in Living Taraxacum kok-saghyz by Ultra-High Performance Liquid Chromatography Tandem Mass Spectrometry

Taraxacum kok-saghyz(TKS)is rich in natural rubber(NR),a natural organic macromolecular compound composed of cis-1,4-polyisoprene,and may become the second NR-bearing plant for biochemical engineering development.In this paper,a rapid and quantitative ultra-high performance liquid chromatography tandem mass spectrometry(UHPLCMS/MS)method was established for determination of macromolecular biosynthesis substrate(dimethylallyl pyrophosphate,DMAPP)and initiator(farnesyl pyrophosphate,FPP)contained in TKS.A Kromasil C18 chromatographic column was used for separation,and the multi-reaction monitoring mode(MRM)of triple quadrupole mass spectrometry was used for detection.Quantification was performed by external calibration method.The results showed that the limit of detection(LOD)and the limit of quantitation(LOQ)of DMAPP were 2.42μg/L and 7.26μg/L,respectively,and the LOQ and the LOD of FPP were 1.02μg/L and 3.05μg/L,respectively.At a concentration of 1—1000μg/L,both analytes had good determination coefficients(>0.999)of calibration curve.The recoveries of DMAPP and FPP were between 99.0%and 117.1%.In real samples detection,the contents of DMAPP and FPP in TKS samples were between 23.32—82.77μg/L and 12.03—85.67μg/L,respectively.Thus,this approach is a reliable method to quantify DMAPP and FPP in TKS.