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A Highly Sensitive LC-MS/MS method for the quantification of Desmopressin in human plasma

06 Apr 2023

Desmopressin is a synthetic analog of vasopressin used to reduce renal excretion of water in central diabetes insipidus and nocturia.
It has been employed clinically since 1972 and is available in various formulations including intranasal solution, intravenous solution, oral tablet and oral lyophilisate. The active ingredient in most formulations is desmopressin acetate. The absolute bioavailability of orally administered desmopressin varies between 0.08% and 0.16%.Because of the low bioavailability, peak plasma concentration of desmopressin after oral administration are very low. Hence highly sensitive method at an LLOQ of 1 pg/mL is required for proper characterization of pharmacokinetic profile. Further, as desmopressin is a synthetic octapeptide, it has been always a challenge to detect such peptides in human plasma at lower detection level (Picogram level). We, at Lambda have developed and validated a highly sensitive and reproducible method of Desmopressin at an LLOQ of 1 pg/mL. The validated method has linearity range of 1 pg/mL to 100 pg/mL

Extraction Method:
Briefly, the method for the quantification of Desmopressin involves solid phase extraction using isotope labeled internal standard (ISTD). Chromatography was achieved on HILIC column with a mobile phase consisting of ammonium acetate and Acetonitrile, with total run-time of 11.5 minute.

This new method was validated in terms of linearity, stabilities, carry-over effect, reinjection reproducibility, precision, accuracy and selectivity. The linearity ranged from 1 to 100 pg/mL. The method exhibited very good precision and accuracy at all concentration levels.

This validated method has been successfully applied to the analysis of study samples in which incurred sample reproducibility experiment was performed and found well within the acceptable criteria which proves method reproducibility for incurred samples.

LC-MS/MS condition:
Molecular weight of Desmopressin is 1069.22 Dalton. Detection was achieved on AB Sciex 6500 LC-MS/MS system using electro spray in positive mode. For this small peptide we choose doubly charged signal as parent ion for better ionization in ion source. MRM transition were measured using 535.5/328.1. In which precursor ion 535.5 amu is related to double charged Desmopressin while its corresponding product ion at 328.1 amu is identified as single charged Gly-Arg-Pro.

Find below global statistics of three precision and accuracy run.

We have generated the matrix stabilities such as Bench top stability for 16.0 hrs, Freeze thaw stabilities for five cycles and long term stability in matrix for 59 days during the course of method validation.

Conclusion: A selective, precise, accurate and highly sensitive LC-MS/MS method is now available for the quantification of Desmopressin in human plasma.

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