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Development and optimization of a gas chromatography/mass spectrometry method for the analysis of thermochemolytic degradation products of phthiocerol dimycocerosate waxes found in Mycobacterium tuberculosis.

Nicoara SC, Minnikin DE, Lee OC, O'Sullivan DM, McNerney R, Pillinger CT, Wright IP, Morgan GH - Rapid Commun. Mass Spectrom. (2013)

Bottom Line: The extraction of PDIMs from culture is simple, and their thermochemolysis is carried out automatically online, thus avoiding the time-consuming derivatization steps of hydrolysis and esterification, usually performed offline.For standard PDIMs in petroleum ether, our optimized method gave an excellent linearity (R(2) = 0.99) at concentrations between 0.172 and 27.5 ng/mL, a good precision (RSD = 11.42%), and a limit of detection (LOD) of 100 pg/mL.For the PDIMs extracted from dilutions of M. tuberculosis culture, the method gave good linearity (R(2) = 0.9685) and an estimated LOD of 400 CFU/mL (CFU = colony forming units) in sterile distilled water.

View Article: PubMed Central - PubMed

Affiliation: Department of Physical Sciences, Faculty of Science, The Open University, Walton Hall, Milton Keynes, MK7 6AA, UK.

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Related in: MedlinePlus

Analysis of the di-methylated phthiocerol A, components C34 and C36, from the thermochemolysis of 6.8 ng of PDIMs: (a) Extracted ion chromatograms showing the fragment ions m/z 201 (gray trace), m/z 339 and m/z 367 (black traces). The mass spectra of these low volatility compounds are shown in (b) and (c). Thermochemolysis conditions: start temperature 70 °C, final PTV temperature 380 °C, temperature ramp rate 4 °C/s, sample transfer time 6.5 min, He carrier gas flow rate 1.4 mL/min. The gas chromatograph oven temperature: 50 °C (9 min) at 30 °C/min to 350 °C (2 min).
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fig04: Analysis of the di-methylated phthiocerol A, components C34 and C36, from the thermochemolysis of 6.8 ng of PDIMs: (a) Extracted ion chromatograms showing the fragment ions m/z 201 (gray trace), m/z 339 and m/z 367 (black traces). The mass spectra of these low volatility compounds are shown in (b) and (c). Thermochemolysis conditions: start temperature 70 °C, final PTV temperature 380 °C, temperature ramp rate 4 °C/s, sample transfer time 6.5 min, He carrier gas flow rate 1.4 mL/min. The gas chromatograph oven temperature: 50 °C (9 min) at 30 °C/min to 350 °C (2 min).

Mentions: The extracted ion chromatograms generated for the methylated phthiocerol A components from 6.8 ng of PDIMs are illustrated in Fig. 4(a). These di-methylated derivatives of the main phthiocerol A components C34 and C36, elute at high oven temperatures of 348 °C and 350 °C, respectively, which increased the thermal stress for the DB-5MS column used, and also extended the analysis time to 21 min. The chromatographic profiles are shown for the fragment ions at m/z 201 (grey trace) and the higher mass fragment ions of m/z 339 and 367 (black traces). Their 70 eV EI molecular ions, expected at m/z 554 (methylated C34) and 582 (methylated C36), were not observed in the mass spectra.


Development and optimization of a gas chromatography/mass spectrometry method for the analysis of thermochemolytic degradation products of phthiocerol dimycocerosate waxes found in Mycobacterium tuberculosis.

Nicoara SC, Minnikin DE, Lee OC, O'Sullivan DM, McNerney R, Pillinger CT, Wright IP, Morgan GH - Rapid Commun. Mass Spectrom. (2013)

Analysis of the di-methylated phthiocerol A, components C34 and C36, from the thermochemolysis of 6.8 ng of PDIMs: (a) Extracted ion chromatograms showing the fragment ions m/z 201 (gray trace), m/z 339 and m/z 367 (black traces). The mass spectra of these low volatility compounds are shown in (b) and (c). Thermochemolysis conditions: start temperature 70 °C, final PTV temperature 380 °C, temperature ramp rate 4 °C/s, sample transfer time 6.5 min, He carrier gas flow rate 1.4 mL/min. The gas chromatograph oven temperature: 50 °C (9 min) at 30 °C/min to 350 °C (2 min).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3824232&req=5

fig04: Analysis of the di-methylated phthiocerol A, components C34 and C36, from the thermochemolysis of 6.8 ng of PDIMs: (a) Extracted ion chromatograms showing the fragment ions m/z 201 (gray trace), m/z 339 and m/z 367 (black traces). The mass spectra of these low volatility compounds are shown in (b) and (c). Thermochemolysis conditions: start temperature 70 °C, final PTV temperature 380 °C, temperature ramp rate 4 °C/s, sample transfer time 6.5 min, He carrier gas flow rate 1.4 mL/min. The gas chromatograph oven temperature: 50 °C (9 min) at 30 °C/min to 350 °C (2 min).
Mentions: The extracted ion chromatograms generated for the methylated phthiocerol A components from 6.8 ng of PDIMs are illustrated in Fig. 4(a). These di-methylated derivatives of the main phthiocerol A components C34 and C36, elute at high oven temperatures of 348 °C and 350 °C, respectively, which increased the thermal stress for the DB-5MS column used, and also extended the analysis time to 21 min. The chromatographic profiles are shown for the fragment ions at m/z 201 (grey trace) and the higher mass fragment ions of m/z 339 and 367 (black traces). Their 70 eV EI molecular ions, expected at m/z 554 (methylated C34) and 582 (methylated C36), were not observed in the mass spectra.

Bottom Line: The extraction of PDIMs from culture is simple, and their thermochemolysis is carried out automatically online, thus avoiding the time-consuming derivatization steps of hydrolysis and esterification, usually performed offline.For standard PDIMs in petroleum ether, our optimized method gave an excellent linearity (R(2) = 0.99) at concentrations between 0.172 and 27.5 ng/mL, a good precision (RSD = 11.42%), and a limit of detection (LOD) of 100 pg/mL.For the PDIMs extracted from dilutions of M. tuberculosis culture, the method gave good linearity (R(2) = 0.9685) and an estimated LOD of 400 CFU/mL (CFU = colony forming units) in sterile distilled water.

View Article: PubMed Central - PubMed

Affiliation: Department of Physical Sciences, Faculty of Science, The Open University, Walton Hall, Milton Keynes, MK7 6AA, UK.

Show MeSH
Related in: MedlinePlus