Assessment of circulating insulin using liquid chromatography-mass spectrometry during insulin glargine treatment in type 2 diabetes: Implications for estimating insulin sensitivity and ß-cell function.

AIM: To determine the potential impact of the cross-reactivity of insulin glargine U-100 and its metabolites on insulin sensitivity and ß-cell measures in people with type 2 diabetes. MATERIALS AND METHODS: Using liquid chromatography-mass spectrometry (LC-MS), we measured concentrations of endogenous insulin, glargine and its two metabolites (M1 and M2) in fasting and oral glucose tolerance test-stimulated plasma from 19 participants and fasting specimens from another 97 participants 12 months after randomization to receive the insulin glargine. The last dose of glargine was administered before 10:00 PM the night before testing. Insulin was also measured on these specimens using an immunoassay. We used fasting specimens to calculate insulin sensitivity (Homeostatic Model Assessment 2 [HOMA2]-S%; QUICKI index; PREDIM index) and ß-cell function (HOMA2-B%). Using specimens following glucose ingestion, we calculated insulin sensitivity (Matsuda ISI[comp] index) and ß-cell response (insulinogenic index [IGI], and total incremental insulin response [iAUC] insulin/glucose). RESULTS: In plasma, glargine was metabolized to form the M1 and M2 metabolites that were quantifiable by LC-MS; however, the analogue and its metabolites cross-reacted by less than 100% in the insulin immunoassay. This incomplete cross-reactivity resulted in a systematic bias of fasting-based measures. By contrast, because M1 and M2 did not change following glucose ingestion, a bias was not observed for IGI and iAUC insulin/glucose. CONCLUSIONS: Despite glargine metabolites being detected in the insulin immunoassay, dynamic insulin responses can be used to assess ß-cell responsiveness. However, given the cross-reactivity of the glargine metabolites in the insulin immunoassay, fasting-based measures of insulin sensitivity and ß-cell function are biased.

Verification of separate measurement procedures where analytical determinations influence the clinical interpretation of GFR: Iohexol quantitation by HPLC and LC-MS/MS.

BACKGROUND: The glomerular filtration rate (GFR) is monitored clinically to follow renal function of a patient. This is commonly performed using endogenous compounds, which estimate GFR (eGFR). However, several conditions exists which may confound or render the eGFR inaccurate. In such cases, it is appropriate to perform a procedure to directly measure GFR (mGFR). Iohexol plasma disappearance is a procedure to determine mGFR and is typically performed using bolus injection of contrast media followed by timed plasma collections. The iohexol plasma concentrations are referenced to the dose given and the elimination rate of iohexol is reflective of the mGFR. Therefore, analytical bias or interference in the iohexol analytical measurement procedure will directly impact the mGFR result. METHODS: Plasma sample iohexol concentrations were measured using both high performance liquid chromatography-ultraviolet detection (HPLC-UV) and liquid chromatography tandem mass spectrometry (LC-MS/MS) measurement procedures. Results were compared on 50 patients where the mGFR was calculated from the iohexol plasma disappearance on two collection time points. RESULTS: Bland-Altman analysis illustrated <1% mean bias when comparing iohexol concentration determinations from the measurement procedures. Passing-Bablok regression revealed y = 1.028x - 0.9420 (slope 95% CI: 1.011, 1.041; Y-intercept 95% CI: -1.606, -0.1638) when comparing LC-MS/MS to HPLC-UV. CONCLUSIONS: Comparison studies of the LC-MS/MS and HPLC-UV measurement procedures displayed a mean bias of <1% by Bland Altman analysis. Measurement of iohexol by LC-MS/MS and HPLC-UV produced similar results and suggests there should be minimal bias in concentration or computed mGFR solely due to the measurement procedure employed.