Classic Prescription, Kai-Xin-San, Ameliorates Alzheimer's Disease as an Effective Multitarget Treatment: From Neurotransmitter to Protein Signaling Pathway.

Alzheimer's disease (AD) is a widespread neurodegenerative disease caused by complicated disease-causing factors. Unsatisfactorily, curative effects of approved anti-AD drugs were not good enough due to their actions on single-target, which led to desperate requirements for more effective drug therapies involved in multiple pathomechanisms of AD. The anti-AD effect with multiple action targets of Kai-Xin-San (KXS), a classic prescription initially recorded in Bei Ji Qian Jin Yao Fang and applied in the treatment of dementia for thousands of years, was deciphered with modern biological methods in our study. Aß 25-35 and D-gal-induced AD rats and Aß 25-35-induced PC12 cells were applied to establish AD models. KXS could significantly improve cognition impairment by decreasing neurotransmitter loss and enhancing the expression of PI3K/Akt. For the first time, KXS was confirmed to improve the expression of PI3K/Akt by neurotransmitter 5-HT. Thereinto, PI3K/Akt could further inhibit Tau hyperphosphorylation as well as the apoptosis induced by oxidative stress and neuroinflammation. Moreover, all above-mentioned effects were verified and blocked by PI3K inhibitor, LY294002, in Aß 25-35-induced PC12 cells, suggesting the precise regulative role of KXS in the PI3K/Akt pathway. The utilization and mechanism elaboration of KXS have been proposed and dissected in the combination of animal, molecular, and protein strategies. Our results demonstrated that KXS could ameliorate AD by regulating neurotransmitter and PI3K/Akt signal pathway as an effective multitarget treatment so that the potential value of this classic prescription could be explored from a novel perspective.

Integrated strategy based on high-resolution mass spectrometry coupled with multiple data mining techniques for the metabolic profiling of Xanthoceras sorbifolia Bunge husks in rat plasma, urine, and feces.

An integrated strategy based on high-resolution mass spectrometry coupled with multiple data mining techniques was developed to screen the metabolites in rat biological fluids after the oral administration of Xanthoceras sorbifolia Bunge husks. Mass defect filtering, product ion filtering, and neutral loss filtering were applied to detect metabolites from the complex matrix. As a result, 55 metabolites were tentatively identified, among which 45 barrigenol-type triterpenoid metabolites were detected in the feces, and six flavonoids and four coumarins metabolites were in the urine. Moreover, eight prototype constituents in plasma, 36 in urine and 23 in feces were also discovered. Due to the poor bioavailability of barrigenol type triterpenoids, most of them were metabolized by intestinal flora. Phase I metabolic reactions such as deglycosylation, oxidation, demethylation, dehydrogenation, and internal hydrolysis were supposed to be their principal metabolic pathways. Coumarins were found in all the biosamples, whereas flavonoids were mainly in the urine. Unlike the saponins, they were mainly metabolized through phase II metabolic reactions like glucuronidation and sulfonation, which made them eliminated more easily by urine. This work suggested the metabolic profile of X. sorbifolia husks for the first time, which will be very valuable for its further development.

A time-of-flight mass spectrometry based strategy to fast screen triterpenoids in Xanthoceras sorbifolia Bunge husks for bioactive substances against Alzheimer's disease.

Xanthoceras sorbifolia Bunge is a folk medicine in China. Recently, the triterpenoids in its husks have attracted more and more attention for potential prevention against Alzheimer's disease. However, current studies on its bioactive substances were still insufficient. To reveal more bioactive substances, an efficient and practical strategy based on high resolution mass spectra coupled with multiple data mining techniques was developed to characterize the barrigenol type triterpenoids in the husks and dosed rat plasma. A total of 50 barrigenol type triterpenoids were identified in the husks, and 6 of these were detected in the rat plasma, which were regarded as bioactive candidates. To find the real bioactive substances, the neuroprotective effect of the candidates was further tested by calculating the PC12 cell viability against amyloid-ß-induced cytotoxicity. As a result, three out of the six candidates exhibited obvious neuroprotction against amyloid-ß-induced cytotoxicity on PC12 cells, indicating their potential to be bioactive substances against Alzheimer's disease. This study will be a valuable reference of the bioactive substances in Xanthoceras sorbifolia Bunge husks against Alzheimer's disease and the provided strategy can also be applied to the exploration of the effective constituents in other medicines.

Metabolomics Strategy Using High Resolution Mass Spectrometry Reveals Novel Biomarkers and Pain-Relief Effect of Traditional Chinese Medicine Prescription Wu-Zhu-Yu Decoction Acting on Headache Modelling Rats.

Headache is a common episodic or chronic neurologic disorder. Treatment options and diagnosis are restricted by an incomplete understanding of disease pathology and the lack of diagnostic markers. Wu-Zhu-Yu decoction (WZYD), a traditional Chinese medicine (TCM) formula containing four TCM herbs, is commonly used in the treatment of headache in China. To deeply understand more about headache and investigate the pain-relief mechanism of WZYD, a comprehensive metabolomics study combined with multivariate data processing strategy was carried out. An LC-high resolution mass spectrometry-based metabolomics approach was applied to characterize metabolic biomarker candidates. Multiple pattern recognition including principal component analysis-discriminant analysis, partial least squares-discriminant analysis and hierarchical cluster analysis were used to determine groups and confirm important variables. A total of 17 potential biomarkers were characterized and related metabolic pathways were identified. The study demonstrated that the established metabolomics strategy is a powerful approach for investigating the mechanism of headache attack and WZYD. In addition, the approach may highlight biomarkers and metabolic pathways and can capture subtle metabolite changes from headache, which may lead to an improved mechanism understanding of central nervous system diseases and TCM treatment.

Development of an ultra-fast liquid chromatography-tandem mass spectrometry method for simultaneous determination of seven flavonoids in rat plasma: Application to a comparative pharmacokinetic investigation of Ginkgo biloba extract and single pure ginkgo flavonoids after oral administration.

For deeper pharmacokinetic investigation and further curative application of ginkgo flavonoids, a delicate, efficient and precise UFLC-MS/MS technique for synchronous quantitation of seven flavonoids, apigenin, luteolin, naringenin, quercetin, diosmetin, kaempferol and isorhamnetin in rat plasma has been established. After mixing with the internal standard (IS) linarin, bio-samples were pretreated via ethyl acetate for liquid-liquid extraction, then isolated at 0.2ml/min flow rate on a Venusil MP C18 chromatographic column (100mm×2.1mm, 3µm) by means of gradient elution. 0.1% formic acid-water and methanol system was used as the mobile phase. Mass spectrometric inspection was conducted on a 4000Q UFLC-MS/MS system with turbo ion spray source in patterns of negative ion and multiple reaction-monitoring (MRM). All calibration curves proved favorable linearity (R2≥0.9918) in linear ranges. Intra-day and inter-day precisions didn't exceed 14.0% for all the analytes, and the accuracy was within 6.9%. Extraction recoveries of analytes and IS were less than ±15.0% of nominal concentrations. This method has been under thorough and firm verification for a comparative pharmacokinetic research after gavage between Ginkgo biloba extract and single pure ginkgo flavonoids. The results demonstrated that there're evident pharmacokinetic discrepancies, and possible structural influences were innovatively proposed. Generally, substitution with 3-hydroxylation, a double bond in ring C, ring B methoxylation often confer longer onset period. The existence of ring B catechol group gives rise to faster clearance.

Qualitative screening of absorbed indoloquinazoline alkaloids and their metabolites in rat plasma after the oral administration of Wu-Zhu-Yu decoction by high-resolution mass spectrometry with multiple data mining algorithms.

A rapid and sensitive liquid chromatography with high-resolution mass spectrometry method with multiple data processing algorithms was developed and applied for the metabolite profiling of evodiamine and its analogous alkaloids in rat plasma after the administration of Wu-Zhu-Yu decoction. All samples were purified using hydrophilic-lipophilic balanced solid-phase extraction cartridges and analyzed by a Sciex TripleTOF 5600(+) mass spectrometer with a 35 min liquid chromatography gradient elution. High-resolution full-scan mass spectrometry and information-dependent acquisition tandem mass spectrometry data were analyzed using multiple data processing approaches. The results indicated that the detected eight prototype alkaloids could be metabolized to 58 metabolites through both phase I and phase II reactions. Oxidation was demonstrated to be the principle metabolic pathway of the parent compounds. The study contributes to the understanding of the absorption and metabolism of the alkaloids in Wu-Zhu-Yu decoction and provides a detailed analysis of scientific data.