The protective effect of electroacupuncture on the renal cortex of SHRs: A metabonomic analysis

Hypertension affects multiple organs in the body during the development of the disease. The antihypertensive effect of acupuncture has been confirmed. How the protective effect of electroacupuncture on the renal cortex of the spontaneously hypertensive rat (SHR) is achieved has not yet been determined. The purpose of this study was to understand the impact of electroacupuncture on the blood pressure of SHRs and the impact on metabolites in the renal cortex, looking for potential differential metabolites and then proceeding to the next step of exploratory research. The experimental animals were divided into four groups: control group, model group, electroacupuncture group and losartan potassium group. Electroacupuncture on bilateral Taichong (LR3) and Zusanli (ST36) lasted for 3 weeks, and the renal cortex was collected for metabonomics research. UHPLC–MS was used to analyze the changes in the metabolic spectrum of renal cortex tissue. The results showed that electroacupuncture can significantly reduce the blood pressure of SHRs. A total of 12 metabolites changed significantly in the comparison between each group and the model group.The possible mechanism is that the primary bile acid biosynthesis, bile secretion, tryptophan metabolism and other metabolic pathways affect the renal cortex.     

Synergistic therapeutic effects of Duzhong Jiangya Tablets and amlodipine besylate combination in spontaneously hypertensive rats using 1H-NMR- and MS-based metabolomics

Duzhong Jiangya Tablet (DJT) composed of Eucommia ulmoides Oliv. and several other traditional Chinese medicines is a Chinese herbal compound, which is clinically used to treat hypertension. The aim of this study was to evaluate the antihypertensive effect of DJT and amlodipine besylate (AB) on the synergistic treatment of spontaneously hypertensive rats (SHRs), and to explore its antihypertensive mechanism. The synergistic therapeutic effect of DJT in combination with AB on SHR was studied using two metabolomics methods based on mass spectrum (MS) and nuclear magnetic resonance. Metabolomics analysis of plasma, urine, liver, and kidney and the combination of orthogonal partial least squares discriminant analysis was performed to expose potential biomarkers. Then, the overall metabolic characteristics and related abnormal metabolic pathways in hypertensive rats were constructed. Blood pressure measurements showed that DJT combined with AB has better effects in treating hypertension than it being alone. A total of 30 biomarkers were identified, indicating that hypertension disrupted the balance of multiple metabolic pathways in the body, and that combined administration restored metabolite levels better than their administration alone. The changes of biomarkers revealed the synergistic therapeutic mechanism of DJT combined with AB, which provided a reference for the combination of Chinese and Western medicines.     

Assessing Adherence to Antihypertensive Medication by Means of Dose-Dependent Reference Plasma Concentration Ranges and Ultra-High Performance Liquid Chromatography-Ion Trap Mass Spectrometry Analysis

Poor adherence to antihypertensive drug therapy is a well-recognized problem and can be assessed by mass spectrometry-based analyses of body fluids. However, contrary statements exist whether drug quantification in blood or qualitative screening in urine is more suitable. The present pilot study aimed to further elucidate the power of blood plasma drug concentrations for adherence monitoring by developing and validating a quantification procedure for nine antihypertensive drugs (amlodipine, bisoprolol, candesartan, canrenone, carvedilol, metoprolol, olmesartan, torasemide, and valsartan) in blood plasma using liquid–liquid extraction and an ultra-high-performance liquid chromatography-ion trap mass spectrometry analysis. The procedure should then be used for an adherence assessment and compared with the results of an established qualitative urine screening. Selectivity, carryover, matrix effect, accuracy, precision, dilution integrity, and stability were successfully validated, except for amlodipine. The applicability was demonstrated by analyzing 19 plasma samples containing 28 antihypertensive drugs and comparing the measured concentrations with calculated dose-dependent reference plasma concentration ranges. The interpretation of plasma concentrations was found to be more sophisticated and time-consuming than that of urine screening results, and adherence could not be assessed in two cases (10%) due to measured plasma concentrations below the lower limit of quantification. However, 14 out of 19 subjects were classified as adherent (75%) and three as nonadherent (15%), in contrast to 19 (100%) that were claimed to be adherent based on the results of the qualitative urine screening. Nevertheless, further data is needed to estimate whether plasma quantification is superior in terms of assessing adherence to antihypertensive medication.     

Rapid determination and comparative pharmacokinetics of tetrahydropalmatine in spontaneously hypertensive rats and normotensive rats

A rapid high‐performance liquid chromatographic method was developed and validated for determination of tetrahydropalmatine (THP), an active component of Rhizoma Corydalis, in rat plasma. The samples were prepared using protein precipitation and separated on an Agilent XDB‐C₁₈ column (150 × 4.6 mm, 5 µm) with the mobile phase consisting of methanol–0.1% phosphate acid solution, adjusted with triethylamine to pH 5.5 (65:35). Good linearity was found within 0.10–10.00 µg/mL of THP in rat plasma sample. The intra‐ and inter‐day precision values were less than 10%. The developed method was successfully applied to assess the pharmacokinetics of THP in spontaneously hypertensive rats (SHR) and normotensive rats. After oral administration of a single dose of THP (60 mg/kg), the maximum plasma concentrations were 6.15 ± 2.1 and 7.54 ± 2.9 µg/mL for normotensive rats and SHR, respectively. The mean values of AUC_0–∞ of THP in SHR were 81.44 ± 45.0 µg h/mL, significantly higher (p < 0.05) than in normotensive rats (44.06 ± 19.6 µg h/mL). The t_1/2 and MRT in SHR were much longer than that in healthy Sprague–Dawley rats, indicating slow elimination of THP in SHR. The results indicated that there are some differences in pharmacokinetics of THP in SHR and Sprague–Dawley rats and it is very important to investigate the pharmacokinetic properties of drugs in pathological conditions. Copyright © 2011 John Wiley & Sons, Ltd.     

Study of disulfide reduction and alkyl chloroformate derivatization of plasma sulfur amino acids using gas chromatography-mass spectrometry

Four disulfide-reducing agents, dithiothreitol (DTT), 2,3-dimercaptopropanesulfonate (DMPS), and the newly tested 2-mercaptoethanesulfonate (MESNA) and Tris(hydroxypropyl)phosphine (THP), were investigated in detail for release of sulfur amino acids in human plasma. After protein precipitation with trichloroacetic acid (TCA), the plasma supernatant was treated with methyl, ethyl, or propyl chloroformate via the well-proven derivatization–extraction technique and the products were subjected to gas chromatographic–mass spectrometric (GC–MS) analysis. All the tested agents proved to be rapid and effective reducing agents for the assay of plasma thiols. When compared with DTT, the novel reducing agents DMPS, MESNA, and THP provided much cleaner extracts and improved analytical performance. Quantification of homocysteine, cysteine, and methionine was performed using their deuterated analogues, whereas other analytes were quantified by means of 4-chlorophenylalanine. Precise and reliable assay of all examined analytes was achieved, irrespective of the chloroformate reagent used. Average relative standard deviations at each analyte level were ≤6%, quantification limits were 0.1–0.2 μmol L⁻¹, recoveries were 94–121%, and linearity was over three orders of magnitude (r ² equal to 0.997–0.998). Validation performed with the THP agent and propyl chloroformate derivatization demonstrated the robustness and reliability of this simple sample-preparation methodology.     

人类可溶性环氧化物水解酶抑制剂的研究进展

高血压是现代人的常见疾病.虽然有很多不同机理的降压药在临床使用,但是由于个体的差异性,高血压的治疗越来越倾向于个体化治疗,因此降压药的使用也不仅仅局限于血压的降低.人类可溶性环氧化物水解酶抑制剂最大的优势在于其在降压的同时还具有显著的抗炎作用.详细地阐述了人类可溶性环氧化物水解酶抑制剂从早期的环氧结构类型到第三代脲类结构的发展过程和近期研究进展.     

Antihypertensive Effect of Angiotensin‐Converting Enzyme Inhibitory Peptide Obtained from Hen Ovotransferrin

Angiotensin I‐converting enzyme (ACE) inhibitory peptide was isolated from the hen ovotransferrin hydrolysate using chymotryptic hydrolysis by two steps of reverse‐phase high‐performance liquid chromatography. The amino sequence of this novel peptide was identified as Lys‐Val‐Arg‐Glu‐Gly‐Thr‐Thr‐Tyr that inhibited ACE activity in vitro in a concentration‐dependent manner with an effective concentration (IC₅₀) of 102.8 μM. Also, this inhibition was identified as noncompetitive using the Lineweaver‐Burk plot. Moreover, the antihypertensive action of this novel peptide was investigated by an intravenous injection into spontaneously hypertensive rats (SHR). A dose‐dependent reduction of systolic blood pressure by this peptide was observed after 40 min of treatment and it decreased the blood pressure markedly at the maximal dose (1 nmol/mL/kg). The maximal blood pressure lowering activity of this peptide was calculated as 163% of captopril (10 pmol/mL/kg) that was used as positive control. In conclusion, the obtained data suggests that Lys‐Val‐Arg‐Glu‐Gly‐Thr‐Thr‐Tyr has an ability to inhibit ACE activity and decrease the systolic blood pressure in hypertensive animals.     

Antihypertensive Effect of Galegine from Biebersteinia heterostemon in Rats

The aerial part of Biebersteinia heterostemon Maxim. (Geraniaceae Biebersteiniaceae) known as ming jian na bao in Chinese, has been traditionally used in Tibetan folk medicine for treatment of diabetes and hypertension. The aim of the present study was to evaluate the effects of galegine obtained from an ethanol extract of the entire Biebersteinia heterostemon plant on the rat’s cardiovascular system in order to characterize its contributions as an antihypertensive agent. The antihypertensive effect of galegine was investigated in pentobarbital-anesthetized hypertensive rats at three dose levels based on the LD₅₀ of galegine. Meanwhile a positive control group received dimaprit with the same procedure. Dimaprit infusion induced a significant hypotension which declined by an average margin of 20%. Simultaneously, single administration of galegine at the doses of 2.5, 5, and 10 mg/kg by intraperitoneal injection induced an immediate and dose-dependent decrease in mean arterial blood pressure (MABP) by an average margin of 40% with a rapid increase in heart rate (HR). We demonstrated that galegine is effective in reducing blood pressure in anesthetized hypertensive rats with rapid onset and a dose-related duration of the effects. The results indicate that galegine was the bioactive compound which can be used as a pharmacophore to design new hypertensive agents.     

Affinity capillary electrophoresis coupling with partial filling technique and field‐amplified sample injection for enantioseparation and determination of DL‐tetrahydropalmatine

A novel, simple and sensitive method for the enantioseparation and determination of DL ‐tetrahydropalmatine (DL ‐THP) was developed using ACE in combination with partial filling technique and field‐amplified sample injection. A chiral selector, i.e. BSA, was used for the enantioseparation of DL ‐THP in ACE. Effects of BSA concentration, pH and separation voltage on the effectiveness of the enantiomer separation were evaluated. In an optimal condition, D ‐ and L ‐THP were completely enantio‐separated in less than 9 min by partially filling an electrophoretic capillary with 50 μmol/L BSA (50 mbar, 100 s) and carrying out an electrophoresis with 20 mmol/L phosphate buffer (pH 7.4) at 15 kV. The sensitivity was further improved by making use of field‐amplified sample injection to lower the LOD (defined as S/N=3) down to 6 ng/mL. Real samples were also tested and promising results for the determination of DL ‐THP enantiomers were obtained.     

The Modulation of Arachidonic Acid Metabolism and Blood Pressure-Lowering Effect of Honokiol in Spontaneously Hypertensive Rats

Background: Cardiovascular diseases have consistently been the leading cause of death in the United States over the last two decades, with 30% of the adult American population having hypertension. The metabolites of arachidonic acid (AA) in the kidney play an important role in blood pressure regulation. The present study investigates the antihypertensive effect of honokiol (HON), a naturally occurring polyphenol, and examines its correlation to the modulation of AA metabolism. Methods: Spontaneously hypertensive rats (SHR) were randomly divided into four groups. Treatment groups were administered HON intraperitoneally at concentrations of 5, 20, and 50 mg/kg. Blood pressure was monitored at seven-day intervals. After a total of 3 weeks of treatment, the rats were euthanized and the kidney tissues were collected to examine the activity of the two major enzymes involved in AA metabolism in the kidney, namely cytochrome P450 (CYP)4A and soluble epoxide hydrolase (sEH). Results: Rats treated with HON did not experience the rise in blood pressure observed in the untreated SHR. High-dose HON significantly reduced blood pressure and inhibited the activity and protein expression of the CYP4A enzyme in the rat kidney. The activity of the sEH enzyme in renal cytosol was significantly inhibited by medium and high doses of HON. Conclusion: Our data demonstrate the antihypertensive effect of HON and provide a novel mechanism for its underlying cardioprotective properties.     

HPLC determination and antihypertensive effect of metipamide

Summary Metipamide was monitored during a two month cap period of treatment to determine whether the wholeblood levels estimated by HPLC provide a relevant indicator of the possible accumulation of the drug., We also analysed antihypertensive activity and biochemical changes in blood of twenty hypertonic patients. Result showed that metipamide is an effective, first-line antihypertensive agent, combining statisfactory blood pressure reduction with low frequency of side effects and a simple once-daily dosage regime.     

Therapeutic monitoring of metipamide during antihypertensive therapy

The aim of our study was to monitor metipamide during a two-month period of treatment and to determine whether the whole-blood levels estimated by high-performance liquid chromatography provide a relevant indicator of possible accumulation of the drug. We also analysed antihypertensive activity and biochemical changes in the blood of twenty hypertonic patients. The results of our clinical trial showed that metipamide is an effective first-line antihypertensive agent, in that it combines satisfactory reduction of blood pressure with a low frequency of side-effects and a simple once-daily dosage regime.     

Calorimetric investigation of the adsorption of binary vapour mixtures of tetrahydropyrrol with methanol and cyclopentane on the homogeneous surface

Adsorption isotherms and differential heats of adsorption of tetrahydropyrrol (THP) and cyclopentane CP), as well as THP and methanol mixtures (mole ratio 1∶1, 1∶2 and 1∶4) on the graphitized carbon black Sterling MT surface were determined. The influence of dilution of [THP] on its ability to undergo homomolecular association was tested. Even 1∶4 dilution of THP does not prevent is association. Methanol strongly interacts with THP in the mixed adsorbed layer forming heteromolecular associates.     

Novel bacteriochlorine for high tissue-penetration: photodynamic properties in human biliary tract cancer cells in vitro and in a mouse tumour model

Photodynamic therapy of bile duct cancer using hematoporphyrin derivative (HPD) and laser light of 630 nm wavelength is confined to a tumouricidal tissue penetration of 4 mm, which might be doubled with laser light between 700 and 800 nm. Therefore, we investigated the photosensitising properties of a novel bacteriochlorine, tetrakis-pyridyl-tetrahydroporphyrin tosylat (THP) with high absorption at 763 nm. Two biliary cancer cell lines (BDC, GBC) were incubated with HPD or THP to assess cellular uptake kinetics, dark cytotoxicity, and photodynamic cytotoxicity (laser light exposure 1-20 J/cm2). Tumours grown from BDC cells in subcutaneous tissue of severe combined immunodeficient mice were treated with laser light of 30 J/cm2 after injection of THP. The concentrations that killed 50% of cells in the dark were 680 microg/ml of HPD, but > 6400 microg/ml of THP in BDC cells, and 220 microg/ml of HPD, but 6400 microg/ml of THP in GBC cells. Both cell lines exhibited uptake and retention of THP and photodynamic cytotoxicity (up to 86% cells killed). THP induced tumour-selective phototoxicity in the cholangiocarcinoma model. The novel bacteriochlorine THP exhibits photosensitiser properties in biliary tract cancer cells in vitro and in vivo and could achieve deep tumouricidal tissue penetration due to photoactivation at 763 nm.     

Steroselective determination of trihexyphenidyl using carboxylmethyl-β-cyclodextrin by capillary electrophoresis with field-amplified sample stacking

A simple, sensitive and low-cost method using capillary electrophoresis coupled with field-amplified sample stacking (FASS) technique has been developed for enantioselective separation and quantification of trihexyphenidyl (THP) enantiomers in human serum. In this work, three kinds of modified β-cyclodextrin were tested as chiral selectors in CE. Among the CDs studied, THP enantiomers could only be separated by carboxylmethyl-β-cyclodextrin (CM-β-CD). A systematic study of the parameters (CD concentration and pH value in CE buffer, separation voltage and temperature, composition of sample solvent, injection voltage and time) affecting chiral separation and on-line concentration of THP enantiomers were investigated and optimized. The optimum FASS method provided a sensitivity enhancement of about 490-fold compared with usual hydrodynamic injection. Limits of detection for each enantiomer were in the low ng ml^− 1 concentration range (0.92 ng ml^− 1 or 3.06 nM). The quantification of each THP enantiomer in human serum was performed after serum sample extraction. To validate this CE-FASS method, linear regression analysis, intra and inter-day precision and recovery were determined with satisfying results.     

Syntheses and rearrangements of tris(hydroxymethyl)phosphine and tetrakis(hydroxymethyl)phosphonium salts

Abstract

Tetrakis(hydroxymethyl)phosphonium (HOCH₂)₄P⁺ salts, particularly the chloride (THPC) and sulfate (THPS), are among the most accessible organophosphorus compounds that can be made quantitatively from PH₃ and formaldehyde in aqueous media under ambient conditions. These phosphonium salts are air-stable, and are widely used as reactants in organic syntheses, as well as in textile or oil industries. Synthesis of tris(hydroxymethyl)phosphine (THP), an active component of the salts, is more complicated because direct synthesis from PH₃ and CH₂O requires either pressure and high temperature, at which THP rearranges into bis(hydroxymethyl)methylphosphine oxide, CH₃P(O)(CH₂OH)₂, or a metal catalyst that is usually difficult or impossible to recover. Syntheses of THP based on neutralization of the salts are more convenient but require pH control and additional steps to separate THP from CH₂O. This review summarizes literature data on syntheses and purification of the salts and THP, and side reactions such as thermal rearrangement and oxidation.     

Determination of 3,4-dihydroxyphenylglycol (DHPG) by HPLC with coulometric detection, and correlation with 3-methoxy-4-hydroxyphenylglycol (MHPG) in human plasma

A method for determining plasma 3,4-dihydroxyphenylethyleneglycol (DHPG), a central noradrenaline (NA) metabolite, is described. The method used HPLC with dual coulometric detection set in screen mode of operation. The isolation of DHPG and related catecholamines (noradrenaline, dopamine and dihydroxybenzylamine) was performed on acid washed alumina extracted with 0.2 M HCIO4 containing EDTA (0.2%), and reduced glutathione as stabilizer. A reversed phase column with an eluting system containing 0.025 M citric acid-sodium hydrogen phosphate buffer in the ratio 3:2 (v:v) and 5% methanol was used. The experimental results of plasma DHPG levels compared favourably with the results from the literature, and a positively significant correlation with plasma MHPG was found. This method could be used as an alternative in central NA assessment.     

Simultaneous determination of tetrahydropalmatine and tetrahydroberberine in rat urine using dispersive liquid-liquid microextraction coupled with high-performance liquid chromatography

Dispersive liquid-liquid microextraction (DLLME) coupled with high-performance liquid chromatography (HPLC)-UV detection was applied in rat urine for the extraction and determination of tetrahydropalmatine (THP) and tetrahydroberberine (THB), both active components in Rhizoma corydalis. Various parameters affecting the extraction efficiency, such as the type and volume of extraction and dispersive solvent, pH, etc. were evaluated. Under the optimal conditions (extraction solvent: 37 μL of chloroform, dispersive solvent: 100 μL of methanol, alkaline with 100 μL of 1 mol/L NaOH, and without salt addition), the enrichment factors of THP and THB were more than 30. The extraction recoveries were 69.8-75.8% and 72.7-77.6% for THP and THB in rat urine, respectively. Both THP and THB showed good linearity in the range of 0.025-2.5 μg/mL, and the limit of quantification was 0.025 μg/mL (S/N=10, n=6). The intra-day and inter-day precision of THP and THB were <12.6%. The relative recoveries ranged from 95.5 to 107.4% and 96.8 to 100.9% for THP and THB in rat urine, respectively. The method has been successfully applied to rat urine samples. The results demonstrated that DLLME is a very simple, rapid and efficient method for the extraction and preconcentration of THP and THB from urine samples.     

Quantitative Determination of Adrenaline Hydrochloride Injection and Noradrenaline Bitartrate Injection by a New HPLC Method via Substitute for Reference Substance

An HPLC method for quantitative determination of adrenaline hydrochloride injection and noradrenaline bitartrate injection was established and validated with a substitute for the reference substance.Phenylephrine hydrochloride was selected as the substitute for the reference substance of adrenaline and noradrenaline bitartrate.The correction factor of phenylephrine hydrochloride with respect to the reference substance of adrenaline and noradrenaline bitartrate was determined under defined conditions.Adrenal...     

Tamm-Horsfall glycoprotein enhances PMN phagocytosis by binding to cell surface-expressed lactoferrin and cathepsin G that activates MAP kinase pathway

The molecular basis of polymorphonuclear neutrophil (PMN) phagocytosis-enhancing activity (PEA) by human purified urinary Tamm-Horsfall glyco- protein (THP) has not been elucidated. In this study, we found human THP bound to lactoferrin (LF) and cathepsin G (CG) expressed on the surface of PMN, identified by a proteomic study with MALDI-TOF- LC/LC/mass spectrometric analysis. Pre-incubation of 10% SDS-PAGE electrophoresed PMN lysates with monoclonal anti-LF or anti-CG antibody reduced the binding with THP. To elucidate the signaling pathway of THP on PMN activation, we found THP enhanced ERK1/2 phosphorylation, reduced p38 MAP kinase phosphorylation, but had no effect on DNA binding of the five NF-kB family members in PMN. To further clarify whether the carbohydrate-side chains or protein-core structure in THP molecule is responsible for THP-PEA, THP was cleaved by different degrading enzymes with carbohydrate specificity (neuraminidase and β-galactosidase), protein specificity (V8 protease and proteinase K) or glycoconjugate specificity (carboxylpeptidase Y and O-sialoglycoprotein endopeptidase). We clearly demonstrated that the intact protein-core structure in THP molecule was more important for THP-PEA than carbohydrate-side chains. Putting these results together, we conclude that THP adheres to surface-expressed LF and CG on PMN and transduces signaling via the MAP kinase pathway to enhance PMN phagocytosis.