Methods to increase sensitivity of LC/MS analysis
The present disclosure discusses a method of separating a sample (e.g., pharmaceutical drug, genotoxic impurity, biomarker, and/or biological metabolite) including coating a metallic flow path of a chromatographic system; injecting the sample into the chromatographic system; flowing the sample through the chromatographic system; separating the sample; and analyzing the separated sample using mass spectroscopy. In some examples, the coating applied to the surfaces defining the flow path is non-binding with respect to the sample—and the separated sample. Consequently, the sample does not bind to the low-binding surface of the coating of the flow path. The applied coating can increase the chromatographic peak area for the sample of the chromatographic system.
1. A method of separating a sample comprising:
providing a chromatographic column having a metal flow path with an alkylsilyl coating on fluid exposed surfaces, the metal flow path with the alkylsilyl coating preventing the sample from binding to underlying metal surfaces;
injecting the sample onto the chromatographic column, wherein the sample comprises 2-hydroxyglutaric acid;
flowing the sample through the chromatographic column; and
separating the sample, wherein the alkylsilyl coating on the metal flow path of the chromatographic column increases a chromatographic peak area of the 2-hydroxyglutaric acid in the sample in comparison to an uncoated chromatographic column having fluid exposed metal surfaces by 3.5× or greater.
2. The method of claim 1 , wherein the alkylsilyl coating is applied to a frit within the chromatographic column.
3. The method of claim 1 , wherein the alkylsilyl coating is applied to wetted surfaces located upstream of the chromatographic column.
4. The method of claim 1 , wherein the chromatographic column comprises a mix-mode chromatographic column.
5. The method of claim 1 , wherein the chromatographic column comprises a phenyl-hexyl chromatographic column.
6. The method of claim 1 , wherein the alkylsilyl coating is deposited from a vaporized source of bis (trichlorosilyl) ethane or bis (trimethoxysilyl) ethane.
7. A method of separating a sample comprising:
providing a chromatographic column having a metal flow path with an alkylsilyl coating on fluid exposed surfaces, the metal flow path with the alkylsilyl coating preventing the sample from binding to underlying metal surfaces;
injecting the sample onto the chromatographic column, wherein the sample comprises 3-phosphoglyceric acid;
flowing the sample through the chromatographic column; and
separating the sample, wherein the alkylsilyl coating on the metal flow path of the chromatographic column increases a chromatographic peak area of the 3-phosphoglyceric acid in the sample in comparison to an uncoated chromatographic column having fluid exposed metal surfaces by 5.8× or greater.
8. The method of claim 7 , wherein the alkylsilyl coating is applied to a frit within the chromatographic column.
9. The method of claim 7 , wherein the alkylsilyl coating is applied to wetted surfaces located upstream of the chromatographic column.
10. The method of claim 7 , wherein the chromatographic column comprises a mix-mode chromatographic column.
11. The method of claim 7 , wherein the chromatographic column comprises a phenyl-hexyl chromatographic column.
12. The method of claim 7 , wherein the alkylsilyl coating is deposited from a vaporized source of bis (trichlorosilyl) ethane or bis (trimethoxysilyl) ethane.
13. The method of claim 7 , wherein the alkylsilyl coating on the metal flow path of the chromatographic column increases a chromatographic peak area of the 3-phosphoglyceric acid in the sample in comparison to an uncoated chromatographic column having fluid exposed metal surfaces by 100× or greater.