Clinical phenotypic and genotypic analysis of 5 pediatric patients with β-ketothiolase deficiency / 中华儿科杂志

Objective: To summarize the clinical and genetic characteristics of children with β-ketothiolase deficiency (BKTD). Methods: The clinical characteristics, biochemical, markers detected by tandem mass spectrometry (MS/MS) and gas chromatography-mass spectrometry (GC/MS), as well as the variants in ACAT1 gene among 5 children with BKTD in Children's Hospital of Chongqing Medical University between October 2018 and December 2022 were retrospectively analyzed. Results: The onset age of the disease in 5 patients (4 males and 1 female) ranged from 9.7 to 28.0 months. During the acute phase, severe metabolic acidosis was observed with a pH of 6.9-7.1, as well as hypoglycaemia (2.3-3.4 mmol/L) and positive urinary ketone bodies (+-++++). Blood levels of methylcrotonyl carnitine, methylmalonyl carnitine and malonyl carnitine were 0.03-0.42, 0.34-1.43 and 0.83-3.53 μmol/L respectively and were significantly elevated. Urinary 2-methyl-3-hydroxybutyric acid was 22-202 and 3-hydroxybutyric acid was 4-6 066, both were higher than the normal levels. Methylcrotonylglycine was mild elevated (0-29). The metabolites detected by MS/MS and GC/MS were significantly reduced after treatment. Analysis of ACAT1 gene mutation was performed in 5 children. Most variants were missense (8/9). Four previously unreported variants were identified: c.678G>T (p.Trp226Cys), c.302A>G (p.Gln101Arg), c.627_629dupTGA (p.Asn209_Glu210insAsp) and c.316C>T (p.Gln106Ter), the first 2 variants were predicted to be damaging by SIFT, PolyPhen-2 and Mutation Taster software. c.316C>T (p.Gln106Ter) is a nonsense variant. Conclusions: β-ketothiolase deficiency is relatively rare, lacks specific clinical manifestations, however severe metabolic acidosis, hypoglycemia, and ketosis during the acute onset were consistent findings. Missense mutations in the ACAT1 gene are common genetic causes of β-ketothiolase deficiency.

Development and validation of liquid chromatography-tandem mass spectrometry method to quantify dasatinib in plasma and its application to a pharmacokinetic study

Abstract Dasatinib, a potent oral multi-targeted kinase inhibitor against Src and Bcr-Abl, can decrease inflammatory response in sepsis. A simple and cost-effective method for determination of an effective dose dasatinib was established. This method was validated in human plasma, with the aim of reducing the number of animals used, thus, avoiding ethical problems. Dasatinib and internal standard lopinavir were extracted from 180 uL of plasma using liquid-liquid extraction with methyl tert-butil ether, followed by liquid chromatography coupled to triple quadrupole mass spectrometry in multiple reaction monitoring mode. For the pharmacokinetic study, 1 mg/kg of dasatinib was administered to mice with and without sepsis. The method was linear over the concentration range of 1-98 ng/mL for DAS in mice and human plasma, with r2>0.99 and presented intra- and interday precision within the range of 2.3 - 6.2 and 4.3 - 7.0%, respectively. Further intra- and interday accuracy was within the range of 88.2 - 105.8 and 90.6 - 101.7%, respectively. The mice with sepsis showed AUC0-t = 2076.06 h*ng/mL and Cmax =102.73 ng/mL and mice without sepsis presented AUC0-t = 2128.46 h*ng/mL. Cmax = 164.5 ng/mL. The described analytical method was successfully employed in pharmacokinetic study of DAS in mice.

Metabolic study of trimetazidine using ultra-high performance liquid chromatography-tandem mass spectrometry

Abstract In the present study, the application of ultra-high performance liquid chromatography-tandem mass spectrometry allowed us to study of known-as well as hitherto unknown-trimetazidine (TMZ) metabolites in human urine and to propose their renal excretion profiles. Urine samples from a healthy volunteer were analyzed at baseline and at 0-4 h, 4-8 h, 8-12 h, and 12-24 h after a single dose of TMZ. A dilute-and-shoot procedure was used as sample treatment before separation. Full-scan spectra of possible metabolites were acquired. Additionally, product ion scan spectra of precursor ions of interest were also acquired at two collision energies. Intact TMZ was a major excretion product, with a maximum concentration at 4-8 h after administration. Moreover, five minor metabolites were observed, namely trimetazidine-N-oxide (M1), N-formyl trimetazidine (M2), desmethyl-trimetazidine O-sulfate (M3), desmethyl-trimetazidine O-glucuronide (M4), and desmethyl-trimetazidine-N-oxide-O-glucuronide (M5). Metabolite M5 has not previously been reported. Excretion curves were constructed based on the chromatographic peak areas of specific mass transitions (precursor ion > product ion) related to each of the detected metabolites

Thirteen new peptaibols with antimicrobial activities from Trichoderma sp / 中国天然药物

From the fungus Trichoderma sp., we isolated seven novel 18-residue peptaibols, neoatroviridins E-K (1-7), and six new 14-residue peptaibols, harzianins NPDG J-O (8-13). Additionally, four previously characterized 18-residue peptaibols neoatroviridins A-D (14-17) were also identified. The structural configurations of the newly identified peptaibols (1-13) were determined by comprehensive nuclear magnetic resonance (NMR) and high-resolution electrospray ionization tandem mass spectrometry (HR-ESI-MS/MS) data. Their absolute configurations were further determined using Marfey's method. Notably, compounds 12 and 13 represent the first 14-residue peptaibols containing an acidic amino acid residue. In antimicrobial assessments, all 18-residue peptaibols (1-7, 14-17) exhibited moderate inhibitory activities against Staphylococcus aureus 209P, with minimum inhibitory concentration (MIC) values ranging from 8-32 μg·mL-1. Moreover, compound 9 exhibited moderate inhibitory effect on Candida albicans FIM709, with a MIC value of 16 μg·mL-1.

Biotransformation differences of ginsenoside compound K mediated by the gut microbiota from diabetic patients and healthy subjects / 中国天然药物

Many natural products can be bio-converted by the gut microbiota to influence pertinent efficiency. Ginsenoside compound K (GCK) is a potential anti-type 2 diabetes (T2D) saponin, which is mainly bio-transformed into protopanaxadiol (PPD) by the gut microbiota. Studies have shown that the gut microbiota between diabetic patients and healthy subjects are significantly different. Herein, we aimed to characterize the biotransformation of GCK mediated by the gut microbiota from diabetic patients and healthy subjects. Based on 16S rRNA gene sequencing, the results indicated the bacterial profiles were considerably different between the two groups, especially Alistipes and Parabacteroides that increased in healthy subjects. The quantitative analysis of GCK and PPD showed that gut microbiota from the diabetic patients metabolized GCK slower than healthy subjects through liquid chromatography tandem mass spectrometry (LC-MS/MS). The selected strain A. finegoldii and P. merdae exhibited a different metabolic capability of GCK. In conclusion, the different biotransformation capacity for GCK may impact its anti-diabetic potency.

Targeted trace ingredients coupled with chemometric analysis for consistency evaluation of Panax notoginseng saponins injectable formulations / 中国天然药物

Evaluating the consistency of herb injectable formulations could improve their product quality and clinical safety, particularly concerning the composition and content levels of trace ingredients. Panax notoginseng Saponins Injection (PNSI), widely used in China for treating acute cardiovascular diseases, contains low-abundance (10%-25%) and trace saponins in addition to its five main constituents (notoginsenoside R1, ginsenoside Rg1, ginsenoside Re, ginsenoside Rb1, and ginsenoside Rd). This study aimed to establish a robust analytical method and assess the variability in trace saponin levels within PNSI from different vendors and formulation types. To achieve this, a liquid chromatography-triple quadrupole mass spectrometry (LC-MS/MS) method employing multiple ions monitoring (MIM) was developed. A "post-column valve switching" strategy was implemented to eliminate highly abundant peaks (NR1, Rg1, and Re) at 26 min. A total of 51 saponins in PNSI were quantified or relatively quantified using 18 saponin standards, with digoxin as the internal standard. This study evaluated 119 batches of PNSI from seven vendors, revealing significant variability in trace saponin levels among different vendors and formulation types. These findings highlight the importance of consistent content in low-abundance and trace saponins to ensure product control and clinical safety. Standardization of these ingredients is crucial for maintaining the quality and effectiveness of PNSI in treating acute cardiovascular diseases.

Biochemical properties of Scedosporium aurantiacum extracellular elastase-like protease / 生物工程学报

Extracellular elastase-like protease is one of the key virulence proteases of Scedosporium aurantiacum. To date, little is known about this enzyme in terms of genetic information, structure, properties and virulence mechanism due to the difficulties in purification caused by its low secretion amount, high specific activity, uncompleted genome sequencing and annotation. This work investigated the gene, structure and enzymatic properties of this enzyme. The S. aurantiacum elastase-like protease from the fungal culture supernatant was analyzed through tandem mass spectrometry (MS/MS) approach, illustrating its primary structure. Bioinformatics tools were employed to predict the conserved domain and tertiary structure, the enzymatic properties were also studied. It turned out that S. aurantiacum extracellular elastase-like protease demonstrated well hydrolysis towards elastin and bovine achilles tendon collagen, with Vmax of 18.14 μg/s and 17.57 μg/s respectively, better than fish scale gelatin, with the lowest hydrolysis effect on casein. Its activity towards elastin was lower than that of the elastase from porcine pancreas, with values of Kcat/Km of 3.541 (μg/s) and 4.091 (μg/s), respectively. It was an alkaline protease, with optimal pH 8.2 and temperature 37 oC. Zn2+ promoted the enzymatic activity while Ca2+, Mg2+, Na+, elastatinal and PMSF inhibited its activity. Its sequence was similar to Paecilomyces lilacinus secreted serine protease (PDB Entry: c3f7oB_) with multiple conserved fractions each containing more than 7 amino acids, thus suitable for design of PCR primer. This study increased our knowledge on S. aurantiacum extracellular elastase-like protease in terms of structure and enzymatic properties, and may facilitate later studies on protein expression and virulence mechanism.

Advances of peptide-centric data-independent acquisition analysis algorithms and software tools / 生物工程学报

Data-independent acquisition (DIA) is a high-throughput, unbiased mass spectrometry data acquisition method which has good quantitative reproducibility and is friendly to low-abundance proteins. It becomes the preferred choice for clinical proteomic studies especially for large cohort studies in recent years. The mass-spectrometry (MS)/MS spectra generated by DIA is usually heavily mixed with fragment ion information of multiple peptides, which makes the protein identification and quantification more difficult. Currently, DIA data analysis methods fall into two main categories, namely peptide-centric and spectrum-centric. The peptide-centric strategy is more sensitive for identification and more accurate for quantification. Thus, it has become the mainstream strategy for DIA data analysis, which includes four key steps: building a spectral library, extracting ion chromatogram, feature scoring and statistical quality control. This work reviews the peptide-centric DIA data analysis procedure, introduces the corresponding algorithms and software tools, and summarizes the improvements for the existing algorithms. Finally, the future development directions are discussed.

Expert consensus on pre-treatment of clinical samples by liquid chromatography-tandem mass spectrometry / 中华预防医学杂志

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) combines the advantages of high separation ability of chromatography and high selectivity, specificity and sensitivity of mass spectrometry, making it one of the most vibrant new technologies in the field of clinical testing. However, the analytical performance is often limited by the characteristics of the sample to be measured. Due to the limited anti-contamination capability of the mass spectrometer, biological samples need to be properly pre-processed to effectively improve the detection performance and achieve accurate detection. The main function of pre-treatment is to selectively separate the target analyte from the biological matrix to reduce interference from other matrix components. At the same time, the target analytes can be concentrated and enriched to improve the analytical sensitivity. At present, there are many kinds of clinical sample pre-treatment methods, and several methods are time-consuming and cumbersome, which brings difficulties to laboratory personnel in method selection, development and standardized operation. Therefore, the purpose of this consensus is to provide guidance for the establishment of laboratory methods and facilitate the standardized development of clinical mass spectrometry measurement.

UPLC-MS/MS Method for Detection of Etomidate and Its Metabolite Etomidate Acid Quantity in Blood / 法医学杂志

OBJECTIVES@#To establish a method for the simultaneous quantitative analysis of etomidate and its metabolite etomidate acid in blood, and to discuss its application value in actual cases.@*METHODS@#Acetonitrile precipitate protein method was used, and C18 column was selected. Gradient elution was performed with acetonitrile and 5 mmol/L ammonium acetate within 6 min. Electrospray ionization source in positive ion mode was used. The internal standard etomidate acid-d5 was obtained by etomidate-d5 alkaline hydrolysis reaction. Ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used for quantitative analysis. The methodological verification was conducted.@*RESULTS@#Etomidate and etomidate acid in blood showed good linear relationship in the quantitative linear range (r>0.999), with the lower limit of quantification was 2.5 ng/mL and 7.5 ng/mL, respectively. The accuracy, precision, recovery rate, and matrix effect of the method met the professional verification standards. The practical application results showed that etomidate and etomidate acid could be detected in the blood of the abusers, and their mass concentrations ranged from 17.24 to 379.93 ng/mL.@*CONCLUSIONS@#The method established in this study can simultaneously quantify etomidate and etomidate acid in blood, which is simple and convenient to operate with accuracy. It can meet the detection needs of actual cases and provide technical support for law enforcement to crack down on etomidate abuse.

Proteomic Difference Analysis of Whole Blood and Bloodstains / 法医学杂志

OBJECTIVES@#To study the changes of protein levels in peripheral blood after it dried.@*METHODS@#The proteins from whole blood and bloodstains were detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and normalized by the label-free quantification (LFQ) method. The differential proteins were analyzed by using R 4.2.1 software, limma and edgeR package. The analysis of biological function, signaling pathway and subcellular localization for the differential proteins was then performed.@*RESULTS@#A total of 623 and 596 proteins were detected in whole blood and bloodstains, respectively, of which 31 were statistically significant in the quantitative results, including 10 up-regulated and 21 down-regulated proteins in bloodstains.@*CONCLUSIONS@#The protein abundances in whole blood and bloodstains are highly correlated, and the variation of protein abundances may be related to the changes of endogenous and structural proteins in cells. The application of proteomics technology can assist the screening and identification of protein biomarkers, thereby introducing new biomarkers for forensic research.

Detection and Analysis of Blood Dexmedetomidine in Drug-Facilitated Cases / 法医学杂志

OBJECTIVES@#To establish a simple and rapid qualitative and quantitative detection method of dexmedetomidine in blood.@*METHODS@#Blood was separated on the Allure PFP Propyl liquid chromatography column with isocratic elution after it was precipitated by acetonitrile and filtered. Qualitative and quantitative analysis of dexmedetomidine was performed using positive ion scan mode and multi-reaction monitoring mode.@*RESULTS@#The limit of detection of dexmedetomidine in blood was 0.2 ng/mL and the limit of quantification was 0.5 ng/mL. The linearity of the method was good in the range of 0.5-1 000 ng/mL, and the correlation coefficient was greater than 0.99. The accuracy of the method was 90.34%-112.67% and the extraction recovery was 50.05%-91.08%, with no significant matrix effect.@*CONCLUSIONS@#This method is simple, selective and suitable for the qualitative and quantitative analysis of dexmedetomidine in blood, which can provide a reference for drug-facilitated cases involving dexmedetomidine.

Rapid Determination of Bucinnazine in Blood by UPLC-MS/MS / 法医学杂志

OBJECTIVES@#To establish a rapid method for the analysis of bucinnazine in blood by UPLC-MS/MS and to apply the method to the practical case.@*METHODS@#After the internal standard was added to blood, the protein was precipitated with 900 μL mixed solution (Vacetonitrile∶Vwater=8∶2). After vortex and centrifugation, the protein was measured through 0.22 μm filter membrane. The separation was performed on C18 chromatography column, with acetonitrile and 5 mmol/L ammonium acetate containing 0.1% formic acid aqueous as mobile phase gradient elution at the flow rate of 0.4 mL/min. Multiple reaction monitoring scan was performed in electrospray positive ion mode, quantitative measurement was performed by internal standard method, and methodological verification was carried out.@*RESULTS@#The linear relationship of bucinnazine in blood was good in the range of 0.5-200 μg/L, the correlation coefficient (r) was 0.999 7, the limit of detection was 0.1 μg/L, the limit of quantitation was 0.5 μg/L, and the recovery was 78.3%-83.8% at 1, 10 and 100 μg/L mass concentration levels. The matrix effect was 69.4%-73.8%, the intra-day precision was 1.9%-2.8%, and the inter-day precision was 2.8%-3.2%, the accuracy was 3.1%-3.5%. The stability test results of 1 and 100 μg/L mass concentrations at -25 ℃ showed that the accuracy (bias) of 10 d was less than 4.5%.@*CONCLUSIONS@#This method has the advantages of simple pre-treatment process, fast sample processing speed, high sensitivity of instrument analysis, good stability of content determination and reliable identification results, and can meet the needs of case identification.

Analysis of disease spectrum for abnormal 3-hydroxyisovalerylcarnitine metabolism identified through newborn screening and clinical diagnosis / 中华医学遗传学杂志

OBJECTIVE@#To explore the disease spectrum for abnormal 3-hydroxyisovalerylcarnitine (C5OH) metabolism identified through newborn screening and clinical diagnosis patients and the key points for differential diagnosis so as to raise the awareness of pediatricians for such diseases.@*METHODS@#Clinical data of 85 neonates with abnormal C5OH metabolism identified from February 2004 to January 2022 at Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine were collected. Their clinical manifestations and results of tandem mass spectrometry (MS/MS), gas chromatography mass spectrometry (GC-MS) and genetic testing were retrospectively analyzed.@*RESULTS@#Among the 85 cases, 46 (54.1%) were identified by neonate screening, whilst 39 (45.9%) were clinically diagnosed patients. Five diseases were diagnosed, including 28 cases with multiple carboxylase deficiency (MCD, 32.9%), 29 cases with 3-methylcrotonyl-coenzymeAcarboxylasedeficiency (MCCD, 34.1%), 4 cases with 3-methylglutaconic acid (3-MGA, 4.7%), 7 cases with 3-hydroxy-3-methylglutaric acid (3-HMG, 8.2%), and 17 cases with beta-ketothiolase deficiency (BKD, 20.0%). The disorders were characterized by sudden onset, anorexia, vomiting, diarrhea, abnormal breathing, consciousness disorder, spasm and developmental delay.@*CONCLUSION@#Among newborns with abnormal C5OH metabolism, MCCD is the most common disorder, which was followed by BKD and MCD. For patients with abnormal C5OH metabolism, MCD is the most common, followed by BKD and 3-HMG. C5OH related diseases have great heterogeneity. Combination of blood acylcarnitine levels, urinary organic acid levels and genetic testing based on clinical characteristics can help to attain the diagnosis.

Protective effect and mechanism of Maiwei Yangfei Decoction on pulmonary fibrosis mice based on Nrf2 regulation of oxidative stress / 中国中药杂志

This study explored the effect and mechanism of Maiwei Yangfei Decoction(MWYF) on pulmonary fibrosis(PF) mice. MWYF was prepared, and its main components were detected by ultra-high-performance liquid chromatography-triple quadrupole tandem mass spectrometry(UPLC-MS/MS). Male C57BL/6J mice were randomly divided into a control group, a model group, a pirfenidone(PFD) group, and low-, medium-, and high-dose MWYF groups, with 10 mice in each group. The PF model was induced in mice except for those in the control group by intratracheal instillation of bleomycin(BLM), and model mice were treated with saline or MWYF or PFD by gavage the next day. The water consumption, food intake, hair, and activity of mice were observed daily. The pathological changes in lung tissues were observed by hematoxylin-eosin(HE) staining, Masson staining, and CT scanning. The level of hydroxyproline(HYP) in lung tissues was detected by alkaline hydrolysis. Immunohistochemistry was used to observe the expression of collagen type Ⅲ(COL3) and fibronectin. The mRNA expression levels of α-smooth muscle actin(α-SMA), type Ⅰ collagen α1(COL1α1), COL3, and vimentin were detected by reverse transcription real-time fluorescence quantitative polymerase chain reaction(RT-qPCR). Superoxide dismutase(SOD) and malondialdehyde(MDA) kits were used to detect oxidative stress indicators in lung tissues and serum. The nuclear translocation of nuclear factor E2-related factor 2(Nrf2) protein was detected by immunofluorescence. The protein and mRNA expression levels of Nrf2, catalase(CAT), and heme oxygenase 1(HO-1) in lung tissues were detected by Western blot and RT-qPCR. Twelve chemical components were detected by UPLC-MS/MS. Animal experiments showed that MWYF could improve alveolar inflammation, collagen deposition, and fibrosis in PF mice, increase body weight of mice, and down-regulate the expression of fibrosis indexes such as HYP, α-SMA, COL1α1, COL3, fibronectin, and vimentin in lung tissues. In addition, MWYF could potentiate the activity of SOD in lung tissues and serum of PF mice, up-regulate the expression level of Nrf2, and promote its transfer to the nucleus, up-regulate the levels of downstream antioxidant target genes CAT and HO-1, and then reduce the accumulation of lipid metabolite MDA. In summary, MWYF can significantly improve the pathological damage and fibrosis of lung tissues in PF mice, and its mechanism may be related to the activation of the Nrf2 pathway to regulate oxidative stress.

Excretion of three alkaloids from Simiao Pills in urine, feces, and bile between normal and type 2 diabetic rats / 中国中药杂志

This study investigated the differences in excretion kinetics of three alkaloids and their four metabolites from Simiao Pills in normal and type 2 diabetic rats. The diabetes model was established in rats by injection of streptozotocin, and the alkaloids in urine, feces, and bile of normal and diabetic rats were detected by LC-MS/MS to explore the effect of diabetes on alkaloid excretion of Simiao Pills. The results showed that 72 h after intragastric administration of the extract of Simiao Pills, feces were the main excretion route of alkaloids from Simiao Pills. The total excretion rates of magnoflorine and berberine in normal rats were 4.87% and 56.54%, which decreased to 2.35% and 35.53% in diabetic rats, which had statistical significance(P<0.05). The total excretion rates of phellodendrine, magnoflorine, and berberine in the urine of diabetic rats decreased significantly, which were 53.57%, 60.84%, and 52.78% of those in normal rats, respectively. After 12 h of intragastric administration, the excretion rate of berberine in the bile of diabetic rats increased significantly, which was 253.33% of that of normal rats. In the condition of diabetes, the excretion rate of berberine metabolite, thalifendine significantly decreased in urine and feces, but significantly increased in bile. The total excretion rates of jateorrhizine and palmatine in the urine increased significantly, and t_(1/2) and K_e changed significantly. The results showed that diabetes affected the in vivo process of alkaloids from Simiao Pills, reducing their excretion in the form of prototype drug, affecting the biotransformation of berberine, and ultimately increasing the exposure of alkaloids in vivo, which would be conducive to the hypoglycemic effect of alkaloids. This study provides references for the clinical application and drug development of Simiao Pills in diabetes.

Differences in chemical components in processing of dried ginger-steamed, sand-fried, and rice swill water-bleached Aconiti Lateralis Radix Praeparata pieces in "Jianchang" faction based on UPLC-MS/MS / 中国中药杂志

This study compared the changes in chemical components during the processing of different types of Aconiti Lateralis Radix Praeparata(ALRP) in "Jianchang" faction, i.e., dried ginger-steamed ALRP pieces(Yin-FP), sand-fried ALRP pieces(Yang-FP), and rice swill water-bleached ALRP pieces(DFP), and provided a scientific basis for the mechanism in toxicity reduction and efficacy enhancement from a compositional perspective. Samples were collected during the processing of the three types of ALRP pieces, yielding raw ALRP pieces, water-bleached Yin-FP, ginger juice-moistened Yin-FP, steamed Yin-FP, water-bleached Yang-FP, sand-fried Yang-FP, water-bleached DFP, rice swill water-bleached DFP, and roasted DFP. Aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine, benzoylhypaconine, aconine, mesaconine, hypaconine, salsolinol, fuziline, and higenamine in the extracts were determined by UPLC-MS/MS, and then content analysis and cluster heatmap analysis were performed on 11 sets of samples. During the processing of the three types of ALRP pieces, bleaching significantly reduced the content of 12 alkaloids; steaming, stir-frying, and roasting significantly reduced the content of diester-type alkaloids(aconitine, mesaconitine, and hypaconitine) and significantly increased the content of monoester-type alkaloids(benzoylaconine, benzoylmesaconine, and benzoylhypaconine) and aminoalcohol-type alkaloids(aconine, mesaconine, and hypaconine). During the processing of Yin-FP, the diester-type alkaloids continuously decreased, while the monoester-type and aminoalcohol-type alkaloids showed an initial decrease followed by an increase. During the processing of Yin-FP, Yang-FP, and DFP, the diester-type alkaloids continuously decreased, while the monoester-type and aminoalcohol-type alkaloids showed an initial decrease followed by an increase. Steamed Yin-FP showed a higher increase in content than fried Yang-FP and roasted DFP. Comprehensive analysis of content differences in toxic and therapeutic components in three ALRP pieces suggests that the distinctive processing methods in "Jianchang" faction can indeed achieve detoxification and efficacy enhancement on ALRP. This study provides references for understanding the mechanisms of action of the three processing methods.

PK/PD model of Chuanxiong gel plaster in treatment of rheumatoid arthritis / 中国中药杂志

In this experiment, the PK/PD fitting model of Chuanxiong(Chuanxiong Rhizoma) in the treatment of rheumatoid arthritis was established in the form of acupoint combined with external application gel paste. Firstly, the rheumatoid arthritis model was induced by ovalbumin, and the articular fluid of rabbits was extracted by microdialysis. The pharmacokinetic process of Chuanxiong in rabbit articular fluid was analyzed by UPLC-MS/MS, and the pharmacokinetic model was established. The pharmacodynamic effects of Chuanxiong on inflammatory factors IL-1β, TNF-α, and IL-6 were analyzed by enzyme-linked immunosorbent assay(ELISA). The pharmacodynamic model was established, and the PK/PD model was obtained by fitting the data of pharmacokinetics and pharmacodynamics. The results of pharmacokinetics showed that the concentration of ligustrolide A in the articular cavity by drug administration on classical acupoint Zusanli(ST 36) was higher than that by Yanglingquan(GB 34), which reflected the advantage of typical acupoint, while ligustrazine concentration was higher after administration through Yanglingquan than through Zusanli, which was different from the traditional acupoint theory. The results of pharmacodynamics showed that the drug had lag effect. The PK/PD model was constructed by fitting the data. When IL-1β was taken as the efficacy index, the PK/PD models of Chuanxiong in typical acupoint Zusanli group, atypical acupoint Yanglingquan group, and non-acupoint group were E=115.28C_e/(3 316.72+C_e), E=108.73C_e/(2 993.47+C_e), and E=101.34C_e/(3 028.51+C_e). When TNF-α was taken as the efficacy index, the PK/PD models of Chuanxiong in typical acupoint Zusanli group, atypical acupoint Yanglingquan group, and non-acupoint group were E=68.31C_e/(3 285.16+C_e), E=59.27C_e/(2 919.86+C_e), and E=53.61C_e/(2 862.87+C_e). When IL-6 was taken as the efficacy index, the PK/PD models of Chuanxiong in typical acupoint Zusanli group, atypical acupoint Yanglingquan group, and non-acupoint group were E=59.92C_e/(3 461.17+C_e), E=58.34C_e/(2 723.51+C_e), and E=49.17C_e/(2 862.76+C_e). The parameters showed that there were significant differences in E_(max), EC_(e50) and k_(eo). The analysis of data found that the PK/PD fitting effect of Zusanli, a typical acupoint, was the best, which proved that it was still the best site for drug administration. To sum up, it shows that there may be bidirectional selectivity between drugs and acupoints.

Comprehensive identification of metabolites and metabolic characteristics of luteolin and kaempferol in Simiao Yong'an Decoction in rats by UHPLC-LTQ-Orbitrap MS/MS / 中国中药杂志

Simiao Yong'an Decoction is a classic prescription for treating gangrene. Modern medical evidence has proven that Si-miao Yong'an Decoction has therapeutic effects on atherosclerosis(AS), vascular occlusion angeitides, and hypertension, while its pharmacodynamic mechanism remains unclear. The evidence of network pharmacology, molecular docking, literature review, and our previous study suggests that luteolin and kaempferol are two major flavonoids in Simiao Yong'an Decoction and can inhibit macrophage inflammation and exert anti-AS effects. However, due to lack of the metabolism studies in vivo, little is known about the metabolic characteristics of luteolin and kaempferol. This study employed ultra-performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS/MS) and relevant software to identify the metabolites and metabolic pathways of luteolin and kaempferol in rat plasma, urine, and feces, after oral administration of luteolin and kaempferol, respectively. After the administration of luteolin, 10, 11, and 3 metabolites of luteolin were detected in the plasma, urine, and feces, respectively. After the administration of kaempferol, 9, 3, and 1 metabolites of kaempferol were detected in the plasma, urine, and feces, respectively. The metabolic pathways mainly involved methylation, glucuronidation, and sulfation. This study enriches the knowledge about the pharmacological mechanism of luteolin and kaempferol and supplies a reference for revealing the metabolic process of other flavonoids in Simiao Yong'an Decoction, which is of great significance for elucidating the pharmacological effects and effective substances of this decoction in vivo.