Lasso-Logistic regression model for the identification of serum biomarkers of neurotoxicity induced by strychnos alkaloids.

As a traditional Chinese medicine, strychnos alkaloids have wide effects including antitumor, analgesic, and anti-inflammatory. However, the therapeutic window of strychnos alkaloids is quite narrow due to potential neurotoxicity. Therefore, it is necessary to explore some efficient biomarkers to identify and predict the neurotoxicity induced by strychnos alkaloids and find a therapy to prevent the neurotoxicity of strychnos alkaloids. Based on the previous studies of our research team, 21 endogenous substances related to neurotoxicity were monitored in rats' serum with HPLC-MS/MS and ELISA. Starting from these fundamentals, a Lasso-Logistic regression model was used to select efficient biomarkers from 21 endogenous substances to predict brain injury and verify the neuroprotective effect of peonies. Under the processing of the Lasso-Logistic regression model, 12 biomarkers were identified from 21 endogenous substances to predict the neurotoxicity induced by strychnos alkaloids. At the same time, the neuroprotective effect of peonies was further confirmed by evaluating the level of 12 biomarkers. The results indicated that the development of the Lasso-Logistic regression model would provide a new, simple and efficient method for the prediction and diagnosis of the neurotoxicity induced by strychnos alkaloids.

The 1 H and 13 C NMR chemical shifts of Strychnos alkaloids revisited at the DFT level.

The density functional theory calculation of 1 H and 13 C NMR chemical shifts in a series of ten 10 classically known Strychnos alkaloids with a strychnine skeleton was performed at the PBE0/pcSseg-2//pcseg-2 level. It was found that calculated 1 H and 13 C NMR chemical shifts provided a markedly good correlation with experiment characterized by a mean absolute error of 0.08 ppm in the range of 7 ppm for protons and 1.67 ppm in the range of 150 ppm for carbons, so that a mean absolute percentage error was as small as ~1% in both cases.

Synthesis and evaluation of Strychnos alkaloids as MDR reversal agents for cancer cell eradication.

Natural products represent the fourth generation of multidrug resistance (MDR) reversal agents that resensitize MDR cancer cells overexpressing P-glycoprotein (Pgp) to cytotoxic agents. We have developed an effective synthetic route to prepare various Strychnos alkaloids and their derivatives. Molecular modeling of these alkaloids docked to a homology model of Pgp was employed to optimize ligand-protein interactions and design analogues with increased affinity to Pgp. Moreover, the compounds were evaluated for their (1) binding affinity to Pgp by fluorescence quenching, and (2) MDR reversal activity using a panel of in vitro and cell-based assays and compared to verapamil, a known inhibitor of Pgp activity. Compound 7 revealed the highest affinity to Pgp of all Strychnos congeners (Kd=4.4µM), the strongest inhibition of Pgp ATPase activity, and the strongest MDR reversal effect in two Pgp-expressing cell lines. Altogether, our findings suggest the clinical potential of these synthesized compounds as viable Pgp modulators justifies further investigation.

New route for synthesis of 2-(2,2-dimethoxyethyl)-1,2,3,4,5,6-hexahydro-1,5-methanoazocino[4,3-b]indole and DFT investigation.

Development of efficient sequences for the synthesis of the title compound (2-(2,2-dimethoxyethyl)-1,2,3,4,5,6-hexahydro-1,5-methanoazocino[4,3-b]indole) (7) was described. The title compound was synthesized through several steps starting from phenylhydrazine hydrochloride and dimethyl (R)-2-(3-oxocyclohexyl)malonate. In this route, all synthesized compounds were observed by spectroscopic tools (FT-IR, NMR): Methyl-2-(2,3,4,9-1H-carbazol-2-yl)acetate (3), 2-(2,3,4,9-tetrahydro-1H-carbazol-2-yl)acetic acid (4), N-(2,2-dimethoxyethyl)-2-(2,3,4,9-tetrahydro-1H-carbazol-2-yl)acetamide (5), 2-(2,2-dimethoxyethyl)-1,2,4,5,6,7-hexahydro-3H-1,5-methanoazocino[4,3-b]indol-3-one (6), 2-(2,2-dimethoxyethyl)-2,3,4,5,6,7-hexahydro-1H-1,5-methanoazocino[4,3-b]indole (7). The central step in these syntheses is the dehydrogenative reaction, which constructs the tetracyclic ring system from a much simpler tetracyclic precursor. The six-stable conformers of the compound (7) were used for further calculations such as FT-IR, NMR, NLO, and FMO analyses, performed at the B3LYP/6-311++G(d,p) level. This work revealed that (7) can be a good material to use in the non-linear optical material because its ß tensor is greater ten times than that of the urea.

LC-MS/MS determination and comparative pharmacokinetics of strychnine, brucine and their metabolites in rat plasma after intragastric administration of each monomer and the total alkaloids from Semen Strychni.

A rapid, specific and sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the simultaneous determination of strychnine, brucine, strychnine N-oxide and brucine N-oxide in rat plasma. Plasma samples were pretreated via simple protein precipitation with methanol and ephedrine hydrochloride was used as internal standard. Chromatographic separation was carried out on an ZORBAX Eclipse XDB-C18 column (2.1×150mm, 3.5µm) by gradient elution with methanol and 10mM ammonium acetate (adjusted to pH 4.0 with formic acid). The quantification of the analytes was performed by mass spectrometry with TurboIonSpray ionization (ESI) inlet in the positive ion multiple reaction monitoring (MRM) mode. The results showed that the calibration curve was linear in the concentration range of 0.510∼306.3ngmL(-1) for strychnine, brucine and 0.102∼306.0ngmL(-1) for strychnine N-oxide and brucine N-oxide, respectively. The intra- and inter-day precisions were less than 14.9%, and the accuracy ranged from 89.4 to 113% at three QC levels for the 4 analytes. The validated method was successfully applied to the pharmacokinetic study of strychnine, brucine, strychnine N-oxide and brucine N-oxide in rat plasma after oral administration of each monomer and the total alkaloids from Semen Strychni. After single oral administration of the total alkaloids from Semen Strychni at 4 dose levels, Cmax, AUC0-t of strychnine and brucine increased and were proportional to the oral doses. In comparative pharmacokinetics studies, no significant difference was found between each monomer and the total strychnos alkaloids on the pharmacokinetic parameters such as Cmax and AUC. Mean Cmax and AUC of strychnine and brucine were slight increased in the monomer groups in comparison to the total strychnos alkaloids groups, which suggested that some other alkaloids in the Semen Strychni may decrease the absorption of strychnine and brucine in body.

Dynamic detection of non-protein-bound strychnine and brucine in rabbit muscle and synovial fluid after topical application of total Strychnos alkaloid patches.

Semen Strychni, a known toxic drug in Chinese pharmacopoeia, is notable for its therapeutic effects on local muscle and joint pain. However, oral administration can be risky. Topically administered drugs accumulate in the topical muscles and knee joints without any major increase in plasma levels; only non-protein-bound drugs in the biological fluids of target tissues are effective for therapeutic effects. A sensitive and rapid ultra performance liquid chromatography - mass spectrometry (UPLC-MS) method coupled with a microdialysis technique was developed to determine the non-protein-bound strychnine (Str) and brucine (Bru) in rabbit muscle and synovial fluid microdialysate. The UPLC separation was carried out using a 1.7µm BEH C18 column (50 mm × 2.1 mm) with a mobile phase consisting of methanol: water (29.5:70.5, v/v) with 0.1% formic acid and 20 mM ammonium acetate in water. The method was validated at concentrations ranging from 0.58 ng/ml to 467.20 ng/ml for Str and from 0.42 ng/ml to 422.40 ng/ml for Bru. Intra-day and inter-day accuracy ranged from 99.1% to 103.2% for Str and from 95.8% to 108.8% for Bru with intra-day and inter-day precision within 9.7%. The proposed method was successfully applied to determine non-protein-bound Str and Bru, and the analysates concentration remained stable in rabbit muscle and synovial fluid after topical application of total Strychnos alkaloid patches, which indicated that total Strychnos alkaloid patches could substitute for the traditional oral administration of Semen Strychni.