[New pretreatment method for detecting petroleum hydrocarbons in soil: silica-gel dehydration and cyclohexane extraction].

Oil is a primary source of energy worldwide. However, the use of oil produces large amounts of pollutants, which are detrimental to the environment. The presence of petroleum hydrocarbons in soil is a critical marker of environmental pollution and safety. Rapid on-site detection technology has been broadly used in emergency tracking, offering critical information support for effective reactions to environmental emergencies. Thus, it is expected to play an increasingly critical role in environmental remediation efforts. The current approach for petroleum hydrocarbon detection in soil mainly involves Soxhlet extraction with a combination of solvents, including acetone and n-hexane. The samples are then analyzed after rotary evaporation, dehydration with anhydrous sodium sulfate, and purification using a magnesium silica-type adsorbent. Unfortunately, this approach requires sample analysis to be performed in the laboratory, which is tedious and time consuming, and consumes large amounts of solvents. Moreover, the rotary evaporator is not portable. Therefore, this method is not appropriate for the rapid on-site detection of petroleum hydrocarbons. In this study, a rapid on-site detection method based on silica-gel dehydration and cyclohexane extraction was developed for the extraction and pretreatment of petroleum hydrocarbons (C10-C40) in soil. First, an appropriate amount of silica gel was added to the soil, and the mixture was completely ground to eliminate moisture. Next, petroleum hydrocarbons were extracted with 40 mL of cyclohexane, and the extract was cleaned by Florisil solid-phase extraction (SPE) column elution. Finally, the samples were analyzed by gas chromatography (GC) to evaluate the above method. The silica gel exhibited optimal adsorption properties compared with anhydrous sodium sulfate, calcium oxide, and molecular sieves, with recovery of 87.5%. The effects of different soil water content (5%, 10%, and 20%) and silica gel (1, 3, 5, and 10 times the moisture content) dosage on the extraction of petroleum hydrocarbons were investigated. The recoveries of petroleum hydrocarbons increased from 74.0% to 103.8% after 15 min of invasive extraction (relative standard deviation, RSD, <10.1%) when silica gel amounting to 10 times the moisture content was used. Five types of silica gels with different properties were purchased from four manufacturers, and the effects of these silica gels on the dehydration and extraction efficiency of petroleum hydrocarbons in soil were assessed. The results showed that amorphous silica gel led to low recoveries (<60%), spherical silica gel achieved extraction efficiencies of approximately 70%-90%, and alkaline silica gel produced recoveries with poor precision. Therefore, neutral spherical silica gel was used for further experiments. The fingerprints of petroleum hydrocarbons with different carbon numbers are an important reference for identifying pollution sources. Thus, ensuring good recoveries throughout the entire carbon range is necessary to ensure the accuracy of the fingerprint analysis results. The proposed method showed good recoveries for petroleum hydrocarbons of all carbon numbers (75%-101%). The findings above indicate that the developed method could be an efficient means to extract petroleum hydrocarbons from soil for both total quantity and fingerprint analyses. Compared with standard methods, the proposed method requires lower solvent dosages and features simpler processing steps. Another advantage of this method is that it does not require the use of highly toxic halogenated solvents; thus, it does not contribute to environmental pollution. It can be applied to the laboratory analysis of soil petroleum hydrocarbons and coupled with other rapid on-site detection techniques for soil petroleum hydrocarbons, such as infrared spectroscopy and portable GC. However, because it does not include a concentration process, the developed method exhibits relatively low sensitivity. In the future, we plan to develop a simple and flexible on-site sample-concentration system to further improve various indicators of this method.

Taxol-induced alteration of intracellular amino-acid profile related to human cervical carcinoma HeLa cell death.

The anti-tumor action of Taxol was investigated in the changes of amino-acids involved in tumor cell survival. By tracing the intracellular amino-acid profiles of HeLa cells treated with non-conditioned and three conditioned media (Taxol, L-alanine, and Taxol + L-alanine), it was observed that an alteration of amino-acid metabolism participates in Taxol-induced death of HeLa cells. The contents of 18 out of 21 detected amino-acids are 5-95% and the ones of lysine and methionine are 158 and 117% of the corresponding contents in the control after treatment with Taxol for 24 h, respectively. Addition of L-alanine inhibited cell apoptosis upon Taxol treatment by partially blocking the increase of lysine and methionine and reversing decrease trend of alanine, glycine, and glutamic acid. These results suggest that interference of amino-acid metabolism might be an important mechanism of Taxol cytotoxicity.

Rhizoma coptidis and berberine-induced activation of murine microglia N9 cells.

AIMS OF THE STUDY: To investigate the effect of water extract of Rhizoma coptidis (WEC) and berberine on the activation of murine microglia N9 cells and corresponding mechanism related to mitochondria. MATERIALS AND METHODS: Phagocytic activity of murine microglia N9 cells was measured by neutral red staining method after the cells were treated with various concentrations of WEC and alkaloids for 24h. Flow cytometric analysis was performed to determine the level of intracellular ROS, Ca(2+), and mitochondrial transmembrane potential (Delta psi) after 87 microg/ml of WEC and 12.4 microg/ml of berberine treatment. Global changes of gene expression in WEC- and berberine-treated N9 cells were measured using cDNA microarray. RESULTS: WEC and berberine, but not palmatine and jatrorrhizine, enhanced phagocytic activity of murine N9 cells in a dose-dependent manner. Both of WEC and berberine stimulated free radical generation, enhanced mitochondrial Delta psi and induced gene expression of Ndufab1, Cox6a2 and Atp5a1. However, a more significant phagocytic effect was observed for WEC. WEC, but not berberine, increased intracellular Ca(2+) concentration. The gene expression of Atp5c1 was selectively up-regulated by WEC, while three genes of Uqcrq, Cox8b, and Atp5g2 were induced by berberine. CONCLUSIONS: WEC and berberine activated murine microglia N9 cells by the regulation of mitochondrial function and mitochondria-related signal molecules. The action of WEC is stronger than that of berberine, indicating that the effect of WEC is ascribed partially, but not totally, to berberine.

Functional analysis of cultured neural cells for evaluating cold/cool- and hot/warm-natured Chinese herbs.

Recently, modern scientific research has been required to understand pharmacological basis of traditional Chinese medicine (TCM) theory based on the ancient clinical experience, and to investigate the molecular mechanisms of action of Chinese herbs. Here, 20 Chinese herbs, classified into 4 properties (hot, warm, cold and cool) of TCM, were analyzed for their ability to exhibit antioxidant action, to enhance glucose uptake by murine microglia N9 cells, and to influence neurotransmitter norepinephrine (NE) release from rat pheochromocytoma PC12 cells. We found a generally protective effect of both hot/warm-natured and cold/cool-natured herbs against H(2)O(2)-induced N9 cell death, partially by elevating superoxide dismutase (SOD) activity. Glucose uptake was elevated after treatment with some hot/warm-natured herbs. In addition, most herbs with hot/warm nature tended to stimulate NE release, while such stimulatory effect was not observed in the herbs with cold/cool nature. Two cold/cool-natured herbs, Rhizoma coptidis and Radix scutellariae, even significantly suppressed the release. These results suggest that the distinct abilities of Chinese herbs to regulate neural cell functions appear to be correlated with their natures identified in traditional TCM theory, and may be a useful guide for their utility in neural degenerative diseases.

Hydrolytic kinetics of lithospermic acid B extracted from roots of Salvia miltiorrhiza.

The hydrolytic kinetics of lithospermic acid B (LAB) extracted from the roots of Salvia miltiorrhiza (Chinese herb: danshen) was investigated by using reversed-phase high-performance liquid chromatography (HPLC) with UV-vis detection. The influences of initial drug concentration, pH and temperature on hydrolysis of LAB were studied in aqueous solutions. The results showed that initial concentration of LAB has no effect on the degradation rate at pH 2.0. The hydrolysis followed pseudo-first-order kinetics at 90 degrees C. The log k(obs)-pH profile indicated that the optimal stability range was at pH 2.0-5.0. The rate constant of overall hydrolysis as a function of temperature under the given conditions obeyed the Arrhenius equation. Analysis of the acid-induced degraded solution of LAB by liquid chromatography-mass spectrometry (LC-MS) revealed at least four degradation products [M-H](-) ion at m/z 197, 137, 537 and 537, respectively. Three of these degradation products, i.e. danshensu (DSU), protocatechuic aldehyde (PRO), and lithospermic acid, were further identified by comparing the retention times with standard samples. According to the structure of LAB and its hydrolysis behavior in solution, the other product was proposed to be the isomer of lithospermic acid.

Kinetics and mechanism of degradation of lithospermic acid B in aqueous solution.

The degradation of lithospermic acid B (LAB) was investigated as a function of buffer concentration, pH and temperature. Stability tests were performed using a stability-indicating high-performance liquid chromatography (HPLC) with UV-vis detection. The degradation followed pseudo-first-order kinetics under all experimental conditions. The maximum stability of LAB was observed at pH 2.0. The logk(pH)-pH profile described by specific acid-base catalysis and water molecules agreed with the experimental results. The overall degradation rate constant as a function of the temperature under the given conditions obeyed the Arrhenius equation. The chemical fate of LAB in mild acidic solution was investigated, and nine degradation products were detected and tentatively identified by LC-MS analysis. The primary degradation pathway involving the cleavage of ester bond and ring-opened of benzofuran in the LAB are proposed.

[Analysis of volatile constituents of root and rhizome of Asarum heterotropoides Fr. var. mandshuricum (Maxim.) Kitag. by gas chromatography-mass spectrometry].

The volatiles of root and rhizome of Asarum heterotropoides Fr. var. mandshuricum (Maxzim. ) Kitag. were extracted with n-hexane and analyzed by capillary gas chromatography-flame ionization detection and gas chromatography-mass spectrometry (GC-MS) with the same column HP-5 (30 m x 0.25 mm i.d.). Initial oven temperature was kept at 40 degrees C for 2 min, then raised to 160 degrees C at 3 degrees C /min, immediately raised to 250 degrees C at 6 degrees C /min, and kept 250 degrees C for 23 min. According to GC-MS analysis, 36 compounds were identified. The main components are safrole, methyleugenol, 3,5-dimethoxytoluene, pentadecane, N-isobutyldodecatetraenamide. Furthermore, the analysis results of root and rhizome were compared. The results indicate that the fundamental constituents are nearly the same, but the quantities of volatiles in root are richer than those in rhizome. In addition, 8 compounds are selected as the criteria of quality control, including 3-carene, 1,8-cineole, eucarvone, methyleugenol, safrole, myristicin, pentadecane and N-isobutyldodecatetraenamide.

[Vectorial angle method for evaluating the similarity between two chromatographic fingerprints of Chinese herb].

AIM: To evaluate the similarity between two chromatographic fingerprints automatically with computer. METHODS: Chromatogram can be treated as vector of hyperspace, and the similarity between them can be counted according to vectorial angle formula. This process was performed with software written in Visual Basic 6.0. The two main functions of this software are automatic peak tracking in two fingerprints under the same analytic condition and computing the similarity automatically. RESULTS: The HPLC fingerprints of eleven kinds of Evodia rutaecarpa (Juss.) Benth (a traditional Chinese herb) from different sources were obtained and the similarities were calculated with this software. This method was shown to be a good way to evaluate the similarity between two fingerprints. A sample washed seven times with hot water can be clearly discriminated from other samples of Evodia rutaecarpa (Juss.) Benth with similar results. CONCLUSION: This method is a good way to evaluate the similarity between two fingerprints and is helpful in quality control of traditional Chinese medicine.

[Establishment of comprehensive fingerprint database of traditional Chinese herbs].

Quality control of traditional Chinese herbs by fingerprint is being paid close attention in these years. A database software has been designed and built to store these fingerprints and other relevant information. The use of this database is very simple and convenient for ordinary analysts who have no experience on professional database software. This software could be used under many kinds of operation systems such as Windows 95, 98, 2000 and Windows NT 4.0, 5.0. By now, nearly 30 fingerprints and their relevant information of ten kinds of traditional Chinese herbs have been input into this database, and the number of fingerprints in this base will be continuously increased in the future. This database is helpful for the analysis and quality control of traditional Chinese herbs.