ABSTRACT
Objectives:To discuss the feasibility of Image J in quantitative analysis on thin layer chromatography (TLC) using gallic acid in Galla Chinensis as research object.Methods:Silica gel GF 254 thin-layer plate was used with chloroform-ethyl formate-formic acid (5:5:1) as the developing solvent and the images were taken under ultraviolet light (254 nm). Polyamide film was used with 75% ethanol-glacial acetic acid (50:1) as the developing solvent and 1% ferric chloride ethanol solution as the chromogenic reagent, and the images were taken under sunlight. Images obtained from the above conditions were imported into Image J to analyze and calculate the content of gallic acid in Galla Chinensis by using external standard two-point method. High-performance liquid chromatography (HPLC) was used with a mobile phase of methanol-0.1% phosphoric acid solution (15:85) at a wavelength of 273 nm to determine the gallic acid content in Galla Chinensis. Results:The quantitation limit of gallic acid on silica gel GF 254 thin-layer plate was 0.401 6 μg, the linear range was 2.855 - 9.515 μg ( r2 = 0.996 0), and the average recovery was 105.12% ( RSD=3.48%); the quantitation limit of gallic acid on polyamide film was 0.363 4 μg, the linear range was 1.427 - 4.758 μg ( r2 = 0.991 5), and the average recovery was 103.75% ( RSD=4.60%). The HPLC method had a quantitative limit of 4.42 ng, a linear range of 0.122-0.977 μg ( r2 = 0.999 9), and a recovery rate of 98.30% ( RSD = 1.40%). The accuracy, repeatability and stability of RSD were all <5%. The gallic acid content measured using Image J showed a maximum relative error of 9.30% and a minimum of 1.62% compared to the HPLC results. Conclusions:Image J is feasible for quantitative analysis of TLC and can be used as a complementary method for quality control of Chinese materia medica.
ABSTRACT
Objective To evaluate the measurement uncertainty of the detecting procedure of amino acids and carnitines by the Waters ACQUITY TQD tandem mass spectrometer and PerkinElmer NeoBaseTM non‐derivatized MSMS kit ,and to discuss the meaning of evaluation .Methods According to the method provided by the Medical Laboratories‐Evaluation and Expression of Measurement Uncertainty ,the internal quality control was detected by using two different batch codes of kit for 20 d ,once before and after the routine sample detection on each working day ,the detector were randomly changed ,and then the measurement uncer‐tainty introduced by measurement reproducibility was calculated;the amino acid external assessment quality control data in 20 times of neonatal hereditary metabolic disease tandem mass spectrometry screening provided by the National Center for Clinical Laborato‐ry in 2014 and 2015 ,and the same and 16 times of carnitines external assessment quality control data were performed the statistics , and then the measurement uncertainty introduced by bias was calculated .Next the relative standard uncertainty and the relative ex‐panded uncertainty according to the measurement uncertainty introduced by bias and measurement reproducibility were calculated . The allowed total error indicators of amino acid and carnitines external quality assessment in the neonatal hereditary metabolic dis‐ease tandem mass spectrometry screening by the National Center for Clinical Laboratory were used as the target expanded uncer‐tainty .Results The relative expanded uncertainties of citrulline ,methionine ,phenylalanine ,propionyl carnitine ,octanoyl carnitine , dodecanoyl carnitine ,palmitoyl carnitine and octadecanoyl carnitine were 19 .1% -26 .1% (k=2) ,which were smaller than the tar‐get uncertainty .The relative expanded uncertainties of leucine ,tyrosine ,valine ,free carnitine were 31 .0% -43 .3% (k=2) ,which were greater than the target uncertainty .The uncertainty of isovaleryl carnitine needed to be estimated separately .Conclusion As‐sessing the measurement uncertainty of the detecting procedure of amino acids and carnitines by the non‐derivatized tandem mass spectrometer method can not only provide an opportunity for continuously improving the detection quality ,but also can help the ex‐perimental technique staffs to interpret the test data correctly and the clinician to use the detection reports correctly .