UPLC-MS/MS simultaneous quantification of urinary circadian rhythm hormones and related metabolites: Application to air traffic controllers.

Civil aviation flight crew and civil aviation air traffic controllers are prone to circadian rhythm abnormalities, which can lead to a slew of other maladies. It could endanger people's health and provide a serious threat to the safety of civil aviation flights if it is not appropriately evaluated and addressed. Early detection of rhythm irregularities and prompt treatment for particular populations that are vulnerable to rhythm disorders are crucial for enhancing civil aviation safety. In general, monitoring of the classical circadian rhythm biomarkers (melatonin or cortisol) in plasma or saliva is an effective way to evaluate the rhythm status. Due to the challenging sample procedure and the trauma of plasma, urine sample testing has received an increasing amount of attention. While, urine circadian rhythm biomarkers have seldom been examined, and the relationship between urinary steroid hormones and melatonin is still poorly understood. In most cases, hormones are determined by immunoassays respectively, mainly enzyme-linked immunosorbent assay (ELISA) or radioimmunoassay (RIA). There are also reports describing the liquid chromatography with tandem mass spectrometry (LC-MS/MS) technique as a method of melatonin or few steroid hormones quantification, however, the simultaneous detection of multiple rhythmic hormones in human urine is rarely reported. For the quantification of the rhythmic hormones in human urine, an accurate approach using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was devised in this work. Nine endogenous hormones (melatonin, 6-hydroxymelatonin, 6-sulfatoxymelatonin, cortisol, corticosterone, cortisone, testosterone, epitestosterone and androsterone), in human overnight urine, were quantified after solid phase extraction (SPE). A reverse phase HSS C18 column was used for chromatographic separation with a 9-minute gradient elution and deuterated analogues of each analyte were applied as internal standards. This method was successfully applied to the analysis of 596 overnight urine samples (23:00-9:00) collected from 84 air traffic controllers in the Beijing area during shift work. This study's findings showed a clear correlation not only between melatonin and its metabolites; cortisol-related metabolites, but also between melatonin metabolites and endogenous metabolites upstream and downstream of cortisol, implying that these two categories of hormones can be used as potential biological rhythm indicators to provide circadian rhythm data support for future studies on circadian rhythm disorders.

Identification of characteristic heroin metabolites in urine based on data-mining technology and multivariate statistics analysis combined with a targeted verification approach for distinguishing heroin abusers.

A common phenomenon shows that ingestion of opium poppy shell-containing drugs can result in a "false-positive" urinalysis test result for mandatory or workplace heroin abuse screening. Owing to the short detection window (8 h in urine) of the characteristic heroin metabolite 6-monoacetylmorphine (6-MAM) confirmation or exclusion of heroin abusers still presents major challenges for toxicologists. In this work, we developed an ultra-performance liquid chromatography-time-of-flight mass spectrometry method (UPLC-TOF-MS) with online data acquisition and multiple post-data-mining technologies combined with a multivariate statistical and batch validation analysis workflow to assess the characteristic urine metabolites of heroin abusers. Based on the proposed methods, 28 characteristic metabolites were structurally identified, and their fragmentation patterns and metabolite pathways were also summarized. Correlation analysis was used to investigate the internal relationship and similarities among the identified metabolites, and seven representative metabolites were selected as "Target-metabolites". Multi-batch urine of samples of heroin abusers were certified based on the UPLC-MS/MS method for further validation of the practicability of using this method for routine analysis. Overall, the target-metabolites can be utilized as assistant "biomarkers" in workplace or mandatory drug screenings. This approach encourages further studies on the development of the "false-positive" identification system.

Application of LC-MS-Based Global Metabolomic Profiling Methods to Human Mental Fatigue.

Mental fatigue is characterized by a reduced capacity for work and a loss of capacity to respond to stimulation and is usually accompanied by a feeling of tiredness and drowsiness. Mental fatigue at work is a serious problem and can raise safety concerns especially in the transportation system. It is believed that mental fatigue is a direct or contributing cause of road and air related accidents and incidents. Psychological studies indicate that fatigue results in reduced work efficiency, alertness, and impaired mental performance. However, its underlying biochemical mechanisms are poorly understood. We hypothesized that the human body is an integrated system, and mental fatigue results in changes not only in psychology but also in biochemistry of the human body. These biochemical changes are detectable in metabolites. We employed global metabolomic profiling methods to screen biochemical changes that occur with mental fatigue in air traffic controllers (ATCs) in civil aviation. A total of 45, all male, ATCs (two batches) were recruited as two mental fatigue groups and 23 executive staff acted as a control group for this study. The volunteers' urine samples were collected before and after their work. The samples were analyzed with liquid chromatography/mass spectrometry equipped with a polar, a weak polar, and a nonpolar column, respectively. Three candidate biomarkers were selected on the basis of statistical significance, coefficient of variance, and compared with data of the three groups. The results suggest that urine metabolites may provide a complete new clue from biochemistry to understand, monitor, and manage human mental fatigue.

[Comparative study on quality of Flos Lonicerae between geo-authentic and non-authentic producing areas].

OBJECTIVE: To compare the quality of Flos Lonicerae between different producing areas. METHOD: ICP-AES, UV and HPLC were used to determine the contents of trace elements, chlorogenic acid, total flavonoids, five iridoid glucosides, hederagenin, and oleanolic acid. SAS software system was used to perform data and cluster analyses. RESULT: The results showed that the geo-authentic crude drug was lower in the contents of Cr and Pb but higher in the contents of chlorogenic acid, total flavonoids, five iridoid glucosides, hederagenin, and oleanolic acid than the non-authentic crude drug. CONCLUSION: The geo-authentic crude drug of Flos Lonicerae is better in quality than the non-authentic crude drug based on the modern chemical analyses, which confirms the validity of traditional geo-based classification.

[Study of blending method for the extracts of herbal plants].

The irregularity in herbal plant composition is influenced by multiple factors. As for quality control of traditional Chinese medicine, the most critical challenge is to ensure the dosage content uniformity. This content uniformity can be improved by blending different batches of the extracts of herbal plants. Nonlinear least-squares regression was used to calculate the blending coefficient, which means no great absolute differences allowed for all ingredients. For traditional Chinese medicines, even relatively smaller differences could present to be very important for all the ingredients. The auto-scaling pretreatment was used prior to the calculation of the blending coefficients. The pretreatment buffered the characteristics of individual data for the ingredients in different batches, so an improved auto-scaling pretreatment method was proposed. With the improved auto-scaling pretreatment, the relative. differences decreased after blending different batches of extracts of herbal plants according to the reference samples. And the content uniformity control of the specific ingredients could be achieved by the error control coefficient. In the studies for the extracts of fructus gardeniae, the relative differences of all the ingredients is less than 3% after blending different batches of the extracts. The results showed that nonlinear least-squares regression can be used to calculate the blending coefficient of the herbal plant extracts.

Interaction study between double-stranded DNA and berberine using capillary zone electrophoresis.

Two non-self-complementary 17-mer double-stranded DNA (dsDNA) with four different central base pairs were designed to systematically investigate the binding affinity and sequence specificity of berberine with dsDNA by capillary zone electrophoresis (CZE). The data analysis with the Kenndler model proved only low affinity between dsDNA and berberine and suggested some weak binding preference of berberine for AATT-containing to GGCC-containing dsDNA. The binding constant, Ka, between berberine and dsDNA(AB) was about (1.0 +/- 0.7) x 10(3) M(-1). In addition, the separation of single-stranded DNA (ssDNA) from dsDNA under simple electrophoretic conditions enabled CZE to be a potentially alternative tool to check the extent of DNA annealing, which is usually done by the time-consuming and labor-intensive slab electrophoresis.

[Similarity system theory to evaluate similarity of chromatographic fingerprints of traditional Chinese medicine].

The similarity evaluation of the fingerprints is one of the most important problems in the quality control of the traditional Chinese medicine (TCM). Similarity measures used to evaluate the similarity of the common peaks in the chromatogram of TCM have been discussed. Comparative studies were carried out among correlation coefficient, cosine of the angle and an improved extent similarity method using simulated data and experimental data. Correlation coefficient and cosine of the angle are not sensitive to the differences of the data set. They are still not sensitive to the differences of the data even after normalization. According to the similarity system theory, an improved extent similarity method was proposed. The improved extent similarity is more sensitive to the differences of the data sets than correlation coefficient and cosine of the angle. And the character of the data sets needs not to be changed compared with log-transformation. The improved extent similarity can be used to evaluate the similarity of the chromatographic fingerprints of TCM.

A new method to evaluate the similarity of chromatographic fingerprints: weighted pearson product-moment correlation coefficient.

The Pearson product-moment correlation coefficient is being used to evaluate the similarity of the high-performance liquid chromatographic fingerprints of traditional Chinese medicine (TCM) in China. It is confirmed that a large range of peak areas produced the wrong results. A new algorithm concerning weighted Pearson product-moment correlation coefficient is proposed in this article. The results for both real cases and simulated data sets show that the weighted Pearson product-moment correlation coefficients allow relatively larger differences for large values, smaller differences for small values, and more reliable results than the unweighted Pearson product-moment correlation coefficients. Weight selection depends on the specific scientific problem.

[Analyses on the trace elements of soils in geo-authentic and non-authentic production areas of Flos lonicearae].

OBJECTIVE: To analyse the trace elements presented in the same germplasm of Flos Lonicerae with the soil in different producing areas of crude drugs and to investigate, the relationship between the trace elements and the characteristics of the geo-authentic Flos Lonicerae in the soil and crude drugs. METHOD: The analyses on the trace elements of soil and crude drugs were made by ICP-AES. RESULT: The contents of Sr, K, Na, Mg and Ca were higher in the geo-authentic areas, and the contents of Ca, Sr and Fe were higher, but the Cr and Pb were lower in the geo-authentic Flos Lonicerae. The geo-authentic crude drugs had a strong tendency to accumulate P and Cu. CONCLUSION: There are no direct relationship between the concentrations of trace elements in Flos Lonicerae and those in their corresponding soil. There are good relationship between the absorption and accumulation of Ca and Cr, Co, Na and Fe; Zn and Co, Cr, Mn; Na and Co; Mg and Mn.