Comparison Log
2025-03-23 07:26:56.185856
mwtab Python Library Version: 1.2.5
Source:      https://www.metabolomicsworkbench.org/rest/study/analysis_id/AN005907/mwtab/...
Study ID:    ST003596
Analysis ID: AN005907
Status:      Inconsistent

Sections "SAMPLEPREP" contain missmatched items: {('SAMPLEPREP_SUMMARY', 'For general metabolomics analysis with HILIC-MS method #1 (see MS section), 4 µL serum was mixed with 60 µL -20°C methanol: acetonitrile: water at 40: 40: 20. The sample was vigorously vortexed, and centrifuged at 4°C at 13,000 g for 10 min. 30 µL supernatant was transferred to the HPLC vial from which 5 µL was injected to the LC-MS. For serum glycerol analysis with HILIC-MS method #2, serum glycerol was first enzymatically derivatized into glycerol-3-phosphate before LC-MS analysis. 4 µL serum was mixed with 90 µL freshly made enzyme solution containing 2 U/mL glycerol kinase, 25 mM Tris-HCl (pH 8.0), 50 mM sodium chloride, 10 mM magnesium chloride, and 1.5 mM ATP, and incubated at room temperature for 15 min. The reaction was stopped by the addition of 400 µL methanol. The sample was vortexed, and centrifuged at 16,000 g for 10 min. The supernatant was transferred to a new tube, and dried using SpeedVac at 45 °C for 90 min. 60 µL of methanol:acetonitrile:water at 40:40:20 was added to the residue, vortexed, and centrifuged at 16,000 g for 10 min. 30 µL supernatant was transferred to the HPLC vial, from which 5 µL was injected to the LC-MS. For serum triglyceride analysis with RP-MS method #3, 4 µL serum was mixed with 50 µL extraction buffer of butanol:methanol at 1:1 volume ratio containing 5 mM ammonium formate. The sample was vigorously vortexed, and centrifuged at 4°C at 13,000 g for 10 min. 30 µL supernatant was carefully taken and transferred to glass insert of LC/MS vials for LC/MS analysis. SPLASH® LIPIDOMIX internal standard with serial dilution was spiked in serum for calibration and triglyceride quantification. In the Data(results) section, 0 means non-detectable labeling, and NA refers to missing measurement (we did not measure certain metabolites in certain samples).'), ('SAMPLEPREP_SUMMARY', 'For general metabolomics analysis with HILIC-MS method #1 (see MS section), 4 µL serum was mixed with 60 µL -20°C methanol: acetonitrile: water at 40: 40: 20. The sample was vigorously vortexed, and centrifuged at 4°C at 13,000 g for 10 min. 30 µL supernatant was transferred to the HPLC vial from which 5 µL was injected to the LC-MS. For serum glycerol analysis with HILIC-MS method #2, serum glycerol was first enzymatically derivatized into glycerol-3-phosphate before LC-MS analysis. 4 µL serum was mixed with 90 µL freshly made enzyme solution containing 2 U/mL glycerol kinase, 25 mM Tris-HCl (pH 8.0), 50 mM sodium chloride, 10 mM magnesium chloride, and 1.5 mM ATP, and incubated at room temperature for 15 min. The reaction was stopped by the addition of 400 µL methanol. The sample was vortexed, and centrifuged at 16,000 g for 10 min. The supernatant was transferred to a new tube, and dried using SpeedVac at 45 °C for 90 min. 60 µL of methanol:acetonitrile:water at 40:40:20 was added to the residue, vortexed, and centrifuged at 16,000 g for 10 min. 30 µL supernatant was transferred to the HPLC vial, from which 5 µL was injected to the LC-MS. For serum triglyceride analysis with RP-MS method #3, 4 µL serum was mixed with 50 µL extraction buffer of butanol:methanol at 1:1 volume ratio containing 5 mM ammonium formate. The sample was vigorously vortexed, and centrifuged at 4°C at 13,000 g for 10 min. 30 µL supernatant was carefully taken and transferred to glass insert of LC/MS vials for LC/MS analysis. SPLASH® LIPIDOMIX internal standard with serial dilution was spiked in serum for calibration and triglyceride quantification. In the Data(results) section, "0" means non-detectable labeling, and NA refers to missing measurement (we did not measure certain metabolites in certain samples).')}
Sections "TREATMENT" contain missmatched items: {('TREATMENT_SUMMARY', 'Treatment #1 Infusion of Isotope Tracers with serum metabolites labeling measurement. On the day of tracer infusion, the mouse was transferred to a new cage without food at 9 AM, and fasted for 4 hours before infusion. Right before the start of infusion, the lock solution was drawn and removed from the in-dwelling jugular vein catheter, and the catheter was flushed with 50 µL saline. The vascular access button was then connected to the infusion tether, which was in turn connected with a swivel on a counter-balance arm. This allowed the mouse to move freely around in the cage. The infusion of U-13C-tracer at a small non-perturbing dose started at 2 PM, and lasted for 30 min for fatty acids and 2.5-3 h for all other tracers. The tracer concentration and infusion rates can be found in the Study Design section. At the end of infusion, 40 µL blood was collected by tail snip. For infusion of 13C-lactate, alanine, and glutamine, given their significant labeling difference in the arterial and venous blood, 40 µL arterial blood was sampled from the carotid artery via the pre-implanted catheter, and used for analysis. The blood was collected into a Microvette tube with clotting activator, and was kept on ice during collection. After infusion, the catheter was flushed with 50 µL saline, and then sealed with 10 ~ 20 µL lock solution. The blood was centrifuged at 5,000 ×g for 10 min. 4 µL serum was aliquoted in separate tubes, and stored at -80°C. --- Treatment #2 Hyperinsulinemia Clamp. C57BL/6J mouse (26.5 ± 1.8 g) was transferred to a new cage without food at 9 AM. After fasting for 4 hours, blood was collected via tail snip for basal glycemia and insulin measurement (noted as time point 0, or T0). A mixture of U-13C-glucose (200 mM), unlabeled glucose (2361 mM) and insulin (66.6 mU/mL) was infused at 3 µL/min/mouse. After 2.5 h, blood was collected for serum glucose labeling and insulin measurement (noted as time point T1). Then U-13C palmitate (8 mM) was infused at 6 µL/min/animal for another 30 min (delivered by a second pump), simultaneously with the glucose and insulin infusion (in the Study Design section, it is noted as 13C-glucose and 13C-C16 with 200 and 8 for concentration and 3 and 6 for infusion rate). Blood was collected at the end for serum glucose and palmitate labeling and insulin measurement (time point T2). Insulin was measured by ELISA Kit following the vendor’s protocol. ---- Treatment #3 Measurement of Triglyceride Circulatory Turnover Flux. The serum triglyceride turnover was measured with a pulse-chase strategy. After fasting for 4 hours, BL/6J mouse was infused with 8 mM U-13C-palmitate at 0.25 µL/min/g for 1-2 hours to label serum triglyceride. The infusion was stopped, and blood was collected by tail snip every 15 minutes for one hour to monitor the decay of M+16 labeling L in palmitoyl-containing triglyceride. An exponential curve was fitted with L=Ae^(-kt) for data points from 15 to 60 min. 0-15 min was excluded from modeling for clearance of labeling in free fatty acids. The triglyceride turnover flux R_a was calculated as R_a=k⋅C⋅V, where C is the total triglyceride concentration in serum, and V the serum volume of 49 µL/g (Clemons, 1992).'), ('TREATMENT_SUMMARY', 'Treatment #1 Infusion of Isotope Tracers with serum metabolites labeling measurement. On the day of tracer infusion, the mouse was transferred to a new cage without food at 9 AM, and fasted for 4 hours before infusion. Right before the start of infusion, the lock solution was drawn and removed from the in-dwelling jugular vein catheter, and the catheter was flushed with 50 µL saline. The vascular access button was then connected to the infusion tether, which was in turn connected with a swivel on a counter-balance arm. This allowed the mouse to move freely around in the cage. The infusion of U-13C-tracer at a small non-perturbing dose started at 2 PM, and lasted for 30 min for fatty acids and 2.5-3 h for all other tracers. The tracer concentration and infusion rates can be found in the Study Design section. At the end of infusion, 40 µL blood was collected by tail snip. For infusion of 13C-lactate, alanine, and glutamine, given their significant labeling difference in the arterial and venous blood, 40 µL arterial blood was sampled from the carotid artery via the pre-implanted catheter, and used for analysis. The blood was collected into a Microvette tube with clotting activator, and was kept on ice during collection. After infusion, the catheter was flushed with 50 µL saline, and then sealed with 10 ~ 20 µL lock solution. The blood was centrifuged at 5,000 ×g for 10 min. 4 µL serum was aliquoted in separate tubes, and stored at -80°C. --- Treatment #2 Hyperinsulinemia Clamp. C57BL/6J mouse (26.5 ± 1.8 g) was transferred to a new cage without food at 9 AM. After fasting for 4 hours, blood was collected via tail snip for basal glycemia and insulin measurement (noted as time point 0, or T0). A mixture of U-13C-glucose (200 mM), unlabeled glucose (2361 mM) and insulin (66.6 mU/mL) was infused at 3 µL/min/mouse. After 2.5 h, blood was collected for serum glucose labeling and insulin measurement (noted as time point T1). Then U-13C palmitate (8 mM) was infused at 6 µL/min/animal for another 30 min (delivered by a second pump), simultaneously with the glucose and insulin infusion (in the Study Design section, it is noted as "13C-glucose and 13C-C16" with "200 and 8" for concentration and "3 and 6" for infusion rate). Blood was collected at the end for serum glucose and palmitate labeling and insulin measurement (time point T2). Insulin was measured by ELISA Kit following the vendor’s protocol. ---- Treatment #3 Measurement of Triglyceride Circulatory Turnover Flux. The serum triglyceride turnover was measured with a pulse-chase strategy. After fasting for 4 hours, BL/6J mouse was infused with 8 mM U-13C-palmitate at 0.25 µL/min/g for 1-2 hours to label serum triglyceride. The infusion was stopped, and blood was collected by tail snip every 15 minutes for one hour to monitor the decay of M+16 labeling L in palmitoyl-containing triglyceride. An exponential curve was fitted with L=Ae^(-kt) for data points from 15 to 60 min. 0-15 min was excluded from modeling for clearance of labeling in free fatty acids. The triglyceride turnover flux R_a was calculated as R_a=k⋅C⋅V, where C is the total triglyceride concentration in serum, and V the serum volume of 49 µL/g (Clemons, 1992).')}