LC and LC-MS/MS studies for identification and characterization of new degradation products of ibrutinib and elucidation of their degradation pathway.

Forced degradation/stress degradation studies of ibrutinib drug were done in hydrolytic (acidic, alkaline and neutral), thermal, photolytic and oxidative degradation conditions in different temperature conditions as per International Conference on Harmonization (ICH) guideline Q1A(R2) in order to identify and characterize degradation products (DPs) of ibrutinib. The study revealed that ibrutinib is extremely sensitive to oxidative degradation even at room temperature. The drug substance is highly sensitive to alkaline hydrolysis and susceptible to acidic hydrolysis at 80 °C temperature condition, whereas found stable in neutral, photolytic and thermal stress conditions. Successful separation of ibrutinib and its ten degradation products formed during stress degradation condition were observed using Waters Acquity UPLC C-18 stationary phase (100 mm × 2.1 mm, 1.7 μm) with gradient elution using mobile phase consisting of Eluent-A: ammonium acetate (20 mm, pH-6) and Eluent-B: acetonitrile. The detection was carried out at 215 nm wavelength. Flow rate was set at 0.3 mL/min with injection volume of 5 μL. The drug substance degraded to one degradation product (DP-I) in acidic hydrolysis, five DPs (DP-I, DP-II, DP-V, DP-VIII and DP-IX) in basic hydrolysis and five DPs (DP-III, DP-IV, DP-VI, DP-VII and DP-X) in oxidative degradation condition. A novel and highly sensitive HRMS/MS/TOF method was developed to identify and characterize all the ten DPs formed during stress study. All the DPs were characterized using ESI positive mode. Except DP-I, all the degradation products formed were found to be new degradation impurities and their fragmentation pathways have never been reported earlier. The proposed mechanism and pathway of degradation products of ibrutinib were discussed and outlined.

Mehta L ，Naved T ，Grover P ，Bhardwaj M ，Mukherjee D ... -  《-》

LC and LC-MS/MS studies for identification and characterization of degradation products of d-tubocurarine chloride.

Forced degradation studies of d-tubocurarine (DTC) was carried out in hydrolytic (acidic, alkaline and neutral), thermal, photolytic and oxidative degradation conditions as per International Conference on Harmonization (ICH) guideline Q1A (R2). The present study revealed that DTC is highly sensitive to oxidative degradation conditions even at room temperature whereas the drug was found to be stable in hydrolytic, photolytic and thermal stress conditions. Separation of DTC and its four degradation products (DPs) (DP-I to DP-IV) formed during stress degradation conditions were achieved on Waters Acquity CSH C18 (1.7 µm, 2.1 mm × 100 mm) column using gradient elution with a mobile phase consisting of Eluent-A: 0.1 % Formic acid Eluent-B: acetonitrile achieved successfully. The detection was carried out at 210 nm wavelength and the flow rate was kept at 0.3 mL/min with a 5 μL injection volume. Also, a highly sensitive and robust HRMS/MS/TOF method was established for the identification and characterization of four DPs formed during the stress study. ESI +ve mode was used throughout the study for ionization of all the DPs. The degradation pathway was also established in the study that is never reported earlier.

Grover P ，Maheshwari S ，Nagarajan K ，Mehta L ，Kumar S ，Rastogi C ... -  《-》

LC-MS/MS studies for identification and characterization of new forced degradation products of dabrafenib and establishment of their degradation pathway.

Dabrafenib (Tafinlar) is used for the treatment of patients with BRAF V600 mutation positive unresectable or metastatic melanoma. Forced degradation study of the drug product and drug substance is very much important in drug development and drug discovery to establish the intrinsic stability and understand its behaviors towards different stress conditions. In the current study, compressive stress testing of dabrafenib has been performed as per the recommendation of ICH guidelines to identify and characterize all major degradation products of dabrafenib (DPD) formed. Drug substances were exposed to different stressed conditions as per ICH recommendations. The present study observed that the dabrafenib drug substance is very much sensitive when exposed to oxidative degradation conditions at 80 °C temperature conditions and also sensitive to photolytic degradation conditions. Dabrafenib is stable when treated in acidic, alkaline, neutral and thermal degradation environments as there is no degradation observed in signification percentage under these stressed conditions. The best separation of eight degradation products and dabrafenib drug substance was obtained in Waters BEH (Ethylene Bridge Hybrid) C-18 column (1.7 µm, 100 mm × 2.1 mm) having mobile phase composed of Formic acid (0.1%) and methanol as Eluent A and Eluent B respectively using 225 nm wavelengths. The volume of injection (5 µL) and flow rate (0.3 mL/min) was set throughout the study. Dabrafenib is highly unstable to oxidative stressed conditions as five major degradation products (DPD-II, DPD-III, DPD-IV, DPD-V and DPD-VII) were obtained when exposed to hydrogen peroxide. When dabrafenib is treated under photolytic degradation conditions, three major DPs were formed (DPD-I, DPD-VI and DP-VIII). These DPs were further identified and characterized on sophisticated HRMS/MS/TOF technique for accurate mass measurement. Characterization of all the degradation products was carried out in the ESI positive mode of ionization. The establishment of the degradation pathway of drug substance and fragmentation pathway of DPs were explained in the present study which was never reported in any literature.

Grover P ，Bhardwaj M ，Mehta L 《-》

Liquid chromatography/electrospray ionization tandem mass spectrometry study of repaglinide and its forced degradation products.

Stress stability studies of drugs have been recognized as an essential part of the drug development process. These studies are used to investigate the intrinsic stability of the drugs and for the development of a selective stability indicating assay method (SIAM). Stress testing is also useful for the formulation and packaging development, shelf-life determination and designing of manufacturing processes. As per regulatory guidelines, stress degradation studies and structural characterization should be carried out to establish degradation pathways of the drug, which is essential from both the efficacy and safety point of view. As the stress stability studies of repaglinide have not been reported in the literature, the present study has been undertaken. Repaglinide (RP), an oral anti-diabetic drug, was subjected to hydrolysis (acidic, alkaline and neutral), oxidation, photolysis and thermal stress conditions as per International Conference on Harmonization (ICH) guidelines Q1A (R2). The chromatographic separation of the drug and its degradation products (DPs) was achieved on an Agilent XDB C-18 column using the gradient elution method with a mobile phase consisting of 20 mM ammonium acetate and acetonitrile at flow rate of 1.0 mL min-1 . The DPs were characterized using liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) in combination with accurate mass measurements. The drug degraded under hydrolytic and oxidative stress, while it was stable under thermal and photolytic stress conditions. In total, six DPs were formed and the LC/MS method described here can resolve all DPs from the parent as well as from each other under various stress conditions. To elucidate the structures of DPs, fragmentation of the [M + H]+ ions of RP and its DPs was studied by using LC/ESI-MS/MS combined with accurate mass measurements. The forced degradation of RP carried out as per ICH guidelines results in the formation of six degradation products which have been characterized using LC/MS/MS in combination with accurate mass measurements.

Purna Chander C ，Raju B ，Ramesh M ，Shankar G ，Srinivas R ... -  《-》

Identification and characterization of forced degradation products of sertraline hydrochloride using UPLC, ultra-high-performance liquid Chromatography-Quadrupole-Time of flight mass spectrometry (UHPLC-Q-TOF/MS/MS) and NMR.

The present study focused on the forced degradation behavior of sertraline hydrochloride (SRT), selective serotonin reuptake inhibitor (SSRI). The drug was exposed to different stressed conditions (hydrolytic, oxidative, thermal and photolytic) according to ICH Q1A (R2) guidelines. The study revealed that SRT was stable in hydrolytic (acidic, basic and neutral) and thermal degradation conditions. In contrast, five degradation products (DPs) were formed under oxidative and photolytic degradation conditions. The chromatographic separation of drug substance and its DPs was achieved on an Acquity HSS T3 column (100 × 2.1 mm, 1.7 μ) using 0.1% formic acid and acetonitrile as the mobile phase in gradient mode using a UHPLC-DAD system. The DPs were identified and characterized by high-resolution LC/MS and LC/MS/MS in ESI positive mode. Two DPs (DP-I and DP-II) were formed when SRT was exposed to oxidative degradation conditions. Three DPs formed (DP III-V) when exposed to photolytic degradation conditions. All the five major DPs were isolated using Preparative HPLC. The structures of major DPs formed were further confirmed using NMR technique (1D and 2D). The proposed mechanism for the formation of the SRT DPs via the photolytic/oxidative stress degradation pathway are discussed and outlined.

Grover P ，Bhardwaj M ，Mukherjee D 《-》