Headspace solid-phase microextraction combined with GC×GC-TOFMS for the analysis of volatile compounds of Coptis species rhizomes.

In this study, the investigation of the volatile compounds of dried rhizomes of Coptis chinensis Franch, C. deltoidea C. Y. Cheng et Hsiao, and C. teeta Wall was carried out to complete the chemical composition of these valuable natural products. Volatile profiles were established and compared after headspace solid-phase microextraction (HS-SPME) using a polydimethylsiloxane/divinylbenzene (PDMS/DVB, 65 µm) fibre coupled to comprehensive 2D gas chromatography time-of-flight mass spectrometry (GC×GC-TOFMS). Analyses were performed and compared on two column-phase combinations (non-polar/polar and polar/non-polar). The majority of the identified compounds eluted as well-separated (pure) components as a result of high-resolution capability of the GC×GC method, which significantly reduces co-elution. Therefore, this increases the likelihood that pure mass spectra can be obtained. More than 290 volatile and semi-volatile organic compounds were tentatively characterized by means of GC×GC in tandem with TOFMS detection. Improved result interpretations were obtained in terms of compound classification based on the organized structure of the peaks of structurally related compounds in the GC×GC contour plot. These compounds are distributed over the chemical groups of hydrocarbons, acids, alkenes, alkynes, aldehydes, ketones, alcohols, esters, furans, and terpenoids. Among all the chemical groups, terpenoids present the higher number of identified compounds (44), alkenes (41), and aldehydes and ketones (28). This study completed the volatile phytochemical analysis of the headspace composition of various Coptis species rhizomes, and should serve as a means to identify the difference between the rhizomes and may also be useful to confirm individual species based on their volatile chemical profile.

Evaluation of World Anti-Doping Agency criteria for anabolic agent analysis by using comprehensive two-dimensional gas chromatography-mass spectrometry.

This work presents the validation study of the comprehensive two-dimensional gas chromatography (GC x GC)-time-of-flight mass spectrometry method performance in the analysis of the key World Anti-Doping Agency (WADA) anabolic agents in doping control. The relative abundance ratio, retention time, identification and other method performance criteria have been tested in the GC x GC format to confirm that they comply with those set by WADA. Furthermore, tens of other components were identified with an average similarity of >920 (on the 0-999 scale), including 10 other endogenous sterols, and full mass spectra of 5,000+ compounds were retained. The testosterone/epitestosterone ratio was obtained from the same run. A new dimension in doping analysis has been implemented by addressing separation improvement. Instead of increasing the method sensitivity, which is accompanied by making the detector increasingly "blind" to the matrix (as represented by selected ion monitoring mode, high-resolution mass spectrometry (MS) and tandem MS), the method capabilities have been improved by adding a new "separation" dimension while retaining full mass spectral scan information. Apart from the requirement for the mass spectral domain that a minimum of three diagnostic ions with relative abundance of 5% or higher in the MS spectra, all other WADA criteria are satisfied by GC x GC operation. The minimum of three diagnostic ions arises from the need to add some degree of specificity to the acquired mass spectrometry data; however, under the proposed full MS scan method, the high MS similarity to the reference compounds offers more than the required three diagnostic ions for an unambiguous identification. This should be viewed as an extension of the present criteria to a full-scan MS method.

Comprehensive two-dimensional gas chromatography improves separation and identification of anabolic agents in doping control.

The application of comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GCxGC-TOFMS) for the analysis of six anabolic agents (AAs) in doping control is investigated in this work. A non-polar-polar column configuration with 0.2microm film thickness (d(f)) second dimension ((2)D) column was employed, offering much better spread of the components on (2)D when compared to the alternative 0.1microm d(f)(2)D column. The proposed method was tested on the "key" AA that the World Anti-Doping Agency (WADA) had listed at the low ngmL(-1) levels (clenbuterol, 19-norandrosterone, epimethendiol, 17alpha-methyl-5alpha-androstane-3alpha,17beta-diol, 17alpha-methyl-5beta-androstane-3alpha,17beta-diol and 3'-OH-stanozolol). The compounds were spiked in a blank urine extract obtained by solid-phase extraction, hydrolysis and liquid-liquid extraction; prior to analysis they were converted to the corresponding trimethylsilyl (TMS) derivatives. The limit of detection (LOD) was below or equal to the minimum required performance limit (MRPL) of 2ngmL(-1) defined by WADA, and the correlation coefficient was in the range from 0.995 to 0.999. The method allows choosing an ion from the full mass spectra which shows the least interference from the matrix and/or the best sensitivity; this can only be done if full scan mass spectral data are available. The advantage of GCxGC over classical one-dimensional GC (1D GC), in terms of separation efficiency and sensitivity, is demonstrated on a positive urine control sample at a concentration of 5ngmL(-1). The obtained similarity to the in-house created TOFMS spectra library at this level of concentration was in the range from 822 to 932 (on the scale from 0 to 999). Since full mass spectral information are recorded, the method allows the retro-search of non-target compounds or new "designer steroids", which cannot be detected with established GC-MS methods that use selected ion monitoring (SIM) mode.

Peak modeling approach to accurate assignment of first-dimension retention times in comprehensive two-dimensional chromatography.

Modeling of first-dimension retention of peaks based on modulation phase and period allows reliable prediction of the modulated peak distributions generated in the comprehensive two-dimensional chromatography experiment. By application of the inverse process, it is also possible to use the profile of the modulated peaks (their heights or areas) to predict the shape and parameters of the original input chromatographic band (retention time, standard deviation, area) for the primary column dimension. This allows an accurate derivation of the first-dimension retention time (RSD 0.02%) which is equal to that for the non-modulated experiment, rather than relying upon the retention time of the major modulated peak generated by the modulation process (RSD 0.16%). The latter metric can produce a retention time that differs by at least the modulation period employed in the experiment, which displays a discontinuity in the retention time vs modulation phase plot at the point of the 180 degrees out-of-phase modulation. In contrast, the new procedure proposed here gives a result that is essentially independent of modulation phase and period. This permits an accurate value to be assigned to the first-dimension retention. The proposed metric accounts for the time on the second-dimension, the phase of the distribution, and the hold-up time that the sampled solute is retained in the modulating interface. The approach may also be based on the largest three modulated peaks, rather than all modulated peaks. This simplifies the task of assigning the retention time with little loss of precision in band standard deviation or retention time, provided that these peaks are not all overloaded in the first or second dimension.

Accelerating analysis for metabolomics, drugs and their metabolites in biological samples using multidimensional gas chromatography.

Gas chromatography (GC) with mass spectrometry (MS) is one of the great enabling analytical tools available to the chemical and biochemical analyst for the measurement of volatile and semi-volatile compounds. From the analysis result, it is possible to assess progress in chemical reactions, to monitor environmental pollutants in a wide range of soil, water or air samples, to determine if an athlete or horse trainer has contravened doping laws, or if crude oil has migrated through subsurface rock to a reservoir. Each of these scenarios and samples has an associated implementation method for GC-MS. However, few samples and the associated interpretation of data is as complex or important as biochemical sample analysis for trace drugs or metabolites. Improving the analysis in both the GC and MS domains is a continual search for better separation, selectivity and sensitivity. Multidimensional methods are playing important roles in providing quality data to address the needs of analysts.

Application of comprehensive two-dimensional gas chromatography to sterols analysis.

The applicability of comprehensive two-dimensional gas chromatography (GCxGC) for sterol analysis was investigated by separation and identification of endogenous sterols in standards, and spiked in human urine. The modulation temperature was optimized to achieve the best separation and signal enhancement. The separation pattern of trimethylsilyl (TMS) derivatives of sterols was compared on two complementary column sets. Whilst the BPX5/BPX50 column set offers better overall separation, BPX50/BPX5 provides better peak shape and sensitivity. Comparison of the identification power of GCxGC-TOFMS against both the NIST05 MS library and a laboratory created (in-house) TOFMS library was carried out on a free sterols extract of urine, derivatised and spiked at the World Anti-Doping Agency (WADA) limit of 2 ng mL(-1). The average match quality for 19 analysed sterols on the BPX50/BPX5 column set was 950/1000 when searched against the in-house library; only four were identified against the NIST05 library, at a match threshold of 800. The match quality of GCxGC-TOFMS spectra was superior to that for analysis using 1D GC-TOFMS for sterols spiked in urine at 10 ng mL(-1). An r(2)>0.997 was obtained for the concentration range between 0.25 ng mL(-1) and 10 ng mL(-1) for three selected sterols. The lowest limit of detection (LOD) was obtained for estrone (0.1 ng mL(-1)) and the highest LOD was for 5alpha-androstan-3alpha,11beta-diol-17-one, epitestosterone and cholesteryl butyrate (1 ng mL(-1)), using a match threshold of at least 800 and signal-to-noise ratio of at least 10. TOFMS coupled to GCxGC enabled satisfactory identification of sterols in urine at their LOD. A minimum acceptable match (MAM) criterion for urinary sterols using 2D retention times and TOF mass spectra is introduced. This study shows that GCxGC-TOFMS yields high specificity for steroids derived from urine, with detection limits appropriate for use in doping control.