Separation, Characterization and quantification of Impurities and Identification of stress degradants of Cabazitaxel by RP-HPLC and LC-ESI-MS techniques

 

S. Hemchand¹*, R. Ravi Chandra Babu¹, Mukthinuthalapati Mathrusri Annapurna²

¹GITAM Institute of Science, GITAM (Deemed to be University), Visakhapatnam, India

²Department of Pharmaceutical Analysis & Quality Assurance, GITAM Institute of Pharmacy,

GITAM (Deemed to be University), Visakhapatnam, India

 

ABSTRACT:

Cabazitaxel is a natural taxoid used to treat prostate cancer in advanced stage. The authors have separated and identified seven impurities of Cabazitaxel using RP-HPLC and LC-MS techniques. Gradient mode was chosen for the separation of impurities in presence of Cabazitaxel and the method was validated as per the International Conference on Harmonization guidelines. The impurities were quantified in the present study and the plausible degradation path was also proposed. Stress degradation studies were performed and the degradants obtained were characterized with the help of mass spectra. Shimadzu HPLC prominence coupled with MS Spectrometry (THERMO) with Sunfire C18 column (100x 4.6 mm, 3.5 µm particle size) was used for performing the chromatographic separation with0.05% formic acid and acetonitrile mixture as mobile phase with flow rate 1.0 ml/min.The injection volume 10µL and wavelength was monitored at 220 nm). The method was applied to the pharmaceutical formulation to study the suitability of the method. The method is very much useful for the pharmacokinetic studies as well as the metabolites study in vitro and in vivo of cancer patients.

 

 

INTRODUCTION:

Cabazitaxel is an anti-cancer drug used for the treatment of prostate cancer¹. Cabazitaxel was approved by US FDA² in June 2010. European Medicines Agency³ also approved Cabazitaxel to use in combination with prednisone for the patients who are resistant to docetaxel treatment. Cabazitaxelis 15-[((1R,2S)-1-hydroxy-2-((1,1-dimethylethyloxy) carbonylamino-2-phenyl ethyl) carbonyl oxy]-(1S,2S,4S,7R,9S,10S,12R,15S)-1-hydroxy-9,12-dimethoxy-10,14,17,17-tetramethyl-4-(methylcarbonyloxy)-11-oxo-2-(phenylcarbonyloxy)-6-oxa tetracyclo [11.3.1.0^3,10.0^4,7] heptadec-13-ene and has molecular weight 835.93 g/mol with molecular formula C₄₅H₅₇NO₁₄.It binds and stabilizes tubulin where microtubular inhibition results due to de-polymerization and cell division and thereby arrests the cell cycle.

 

Many researchers had worked on Cabazitaxel previously for its quantitative determination using HPLC^4-10 and mass spectral techniques in human plasma^11-12 and in rat whole blood¹³. At present the authors have proposed a new stability indicating liquid chromatographic (RP-HPLC) method as well as LC-MS technique for the determination of Cabazitaxel along with seven impurities. The authors also characterized the degradants products formed during the stress degradation studies using LC-MS data and compared with the known impurities for confirmation. Shimadzu HPLC prominence coupled with MS Spectrometry (Thermo) with Sunfire C18 column (100 x 4.6 mm, 3.5 µm particle size) was used for the chromatographic study. 0.05% formic acid and acetonitrile mixture was chosen as mobile phase and the system was operated on gradient mode with flow rate 1.0 ml/min.The injection volume was 10µL and detection wavelength was 220 nm. Cabazitaxel was subjected to stress degradation studies and the degradant products were analyzed using mass spectroscopy. Impurities (I - VII) were separated on gradient mode and characterized using mass spectroscopy. Cabazitaxel and its seven impurities were quantified and the method was validated (ICH guidelines) ^14-16. The chemical structures of Cabazitaxel and its seven impurities were given in Figure 1.

 

Cabazitaxel	Impurity I - (2R,3S) N-BOC-Phenyl isoserine
Impurity II (4S,5R) -3-Tert-butoxycarbonyl-2,2-dimethyl-4-phenyl-  5-oxazolidine carboxylic acid	Impurity III – 10-Deacetyl baccatin–III   (1S,2S,4S,7R,9S,10S,12R,15S,)-1,9,12,15-tetrahydroxy- 10,14,17,17-tetramethyl-4-(methylcarbonyloxy)-11)-oxo-2-(phenylcarbonyloxy)-6-oxa tetra cyclo [11.3.1.03,10.04,7] heptadec-13-ene
Impurity IV – Free amine   (2aR,4S,4aS,6R,9S,11S,12S,12bS)-12b-acetoxy-9-(((2R,3S)-3-amino-2-hydroxy-3-phenyl propanoyl) oxy)-11-hydroxy-4,6-dimethoxy-4a,8,13,13-tetramethyl-5-oxo-2a, 3 ,4, 4a, 5, 6, 9, 10, 11, 12, 12a, 12b-dodecahydro-1H-7,11-methano cyclodeca [3,4] benzo [1,2-b]oxet-12-yl benzoate	Impurity V – 7,10-Diethoxy-10-DAB-III   (1S,2S,4S,7R,9S,10S,12R,15S)-1,15-dihydroxy-9,12-dimethoxy-10,14,17,17-tetramethyl-4- (methylcarbonyloxy)-11)-oxo-2-(phenylcarbonyloxy)-6-oxa tetra cyclo [11.3.1.03,10.04,7] heptadec-13-ene
Impurity VI – Coupled product   (4S,5R)-5-((2aR,4S,4aS,6R,9S,11S,12S,12bS)-12b-acetoxy-12-(benzoyloxy)-11-hydroxy-4,6-dimethoxy-4a,8,13,13-tetramethyl-5-oxo-2a,3,4,4a,5,6,9,10,11,12,12a,12b-dodecahydro-1H-7,11-methanocyclodeca[3,4]benzo[1,2-b]oxet-9-yl)3-tert-butyl2,2-dimethyl-4-phenyl oxazolidine-3,5-dicarboxylate	Impurity VII –Docetaxel anhydrous   (2aR,4S,4aS,6R,11S,12S,12bS)-12b-acetoxy-9-(((2R,3R)-3- ((tertbutoxy carbonyl) amino)-2-hydroxy-3-phenyl propanoyl) oxy)-4,6,11-trihydroxy-4a,8,13,13-tetramethyl-5-oxo-2a,3,4,4a,5,6,9, 10, 11,12,12a,12b-dodecahydro-1H-7,11-methano cyclodeca [3,4] benzo[1,2-b]oxet-12-yl benzoate
Figure 1: Chemical structures of Cabazitaxel and its seven Impurities (I-VII)

MATERIALS AND METHODS:

Cabazitaxel API (Active pharmaceutical ingredient) and its seven impurities were procured from Dr. Reddy’s Labs (Hyderabad, India) and used as it is without further purification.HPLC grade acetonitrile, formic acid, hydrochloric acid, sodium hydroxide and 30% w/v hydrogen peroxide were purchased from Merck and Milli-Q water from Millipore system was used.Cabazitaxel stock solution spiked with its seven impurities (I-VII) were prepared in acetonitrile and stored.

 

Chromatographic conditions

Cabazitaxel and its impurities were quantified using Shimadzu HPLC prominence Model coupled with MS Spectrometry (Thermo) with X-Caliber software. Mass detector (Ion trap mass detector) with Electrospray ionization (both positive and negative modes) maintaining capillary temperature 250 °C (Spray/Source voltage (kV): 4.5; Sheath gas flow (Arb): 30.00; Aux/Sweep gas flow (Arb): 5.00). Sunfire C18 column (100 x 4.6 mm, 3.5 µm particle size) was employed and 0.05% formic acid and acetonitrile mixture was used as mobile phase with injection volume 10µL. The system was operated on gradient mode with flow rate 1.0 ml/min monitoring wavelength at 220 nm. Column temperature:25°C and that of the solutions at 15°C.

 

Method validation¹⁴

Series ofCabazitaxel(0 - 300 µg/mL) solutions spiked along with its sevenimpurities (I-VII) were prepared and10 µl of each mixture was injected in to the HPLC system and the system was operated on gradient mode. Thepeak area and thereby the average peak area (n=3) was calculated from the resulting chromatograms and calibration curve wasdrawn using mean peak area on the y-axis and the concentration of the analyte on the x-axis. Precision studies (n=6) were performed for the mixture containing Cabazitaxeland its seven impurities (I-VII) and the % RSD of the mean peak area was determined. Accuracy study was performed for Cabazitaxel and its seven impurities (i.e. 50%, 100% and 150%). Robustness was also performed by varying flow rate (0.2 ml/min), organic phase ratio (5%) and column temperature (5°C).

 

Stress degradation studies¹⁵

Cabazitaxel was exposed to different stress conditions and the fate of the drug was studied with the help of mass spectra.Basic hydrolysis was performed by treating Cabazitaxel with 0.005 N NaOHfor 22 hours at room temperature in dark place where as acidic hydrolysis was performed with 0.5 N HCl for 24 hours at room temperature in dark place. Both acid hydrolyzed and base hydrolyzed drug samples were neutralized before injecting in to the HPLC system. Oxidationwas conducted by treating the drug with 10% H₂O₂for 24 hours at room temperature in dark place. Photolysis was conducted by exposing the drug for 1.2 million lux hours and 200 watt hr/sq. meter and thermal degradation was conducted by heating the drug at 60°c for 9 days in a Petri dish.

 

Experimental application to Cabazitaxel injection

Cabazitaxel injectionis available (60 mg/1.5 ml) with brand names Cabapan (Heet Healthcare Pvt Limited), Jevtana (Dheer Healthcare Ltd; Nexus LifecarePvt Ltd), Kabanat (Shubham Pharmaceuticals). Three different brands were procured, extracted with acetonitrile and then diluted with mobile phase as per the requirement. The percentage purity of Cabazitaxel was determined using the proposed method.

 

RESULTS AND DISCUSSION:

Specific and sensitive stability indicating HPLC andLC-ESI-MSmethods wereestablishedfor the quantification of Cabazitaxel and its seven impurities on gradient mode using formic acid and acetonitrile mixture. The reported methods in the literature were discussed and compared with the presentin Table 1.

 

Table.1. Evaluation of reported liquid chromatographic methods with the present study

Mobile phase (v/v)	λ (nm)	Linearity (mg/ml)	Method	Ref.
Acetonitrile:  phosphate buffer (pH 5.0) (70:30)	230	0.1-150	HPLC	4
Methanol:  0.1% ortho phosphoric acid (80: 20)	210	0.1-200	HPLC	5
Phosphate buffer: Acetonitrile	230	0.025-1.5	Impurities	6
Acetonitrile: sodium acetate buffer (pH 4.0) (30:70)	210	0.1-150	HPLC	7
Acetonitrile: tetra butyl ammonium hydrogen sulphate (70 : 30)	231	0.1–150	HPLC	8
Methanol:  tetra butyl ammonium hydrogen sulphate(70: 30)	210	0.1-200	HPLC	9
Methanol: Sodium acetate buffer (80 : 20)	234	0.1-200	HPLC	10
Ammonium hydroxide: methanol (83:17)	275	2-20	Human plasma LC-MS	11
Acetonitrile: Ammonium formate	362	0.01-0.1	Human plasma LC-MS	12
Acetonitrile: Ammonium acetate (80:20)	236	2.49-99.6	Rat blood (dry blood spots)             (LC-MS/MS)	13
0.05% Formic acid: Acetonitrile (Gradient mode)	220	0-300	HPLC and LC-ESI-MS & Seven Impurities	Present work

 

Method optimization

The RP-HPLC method was optimized using 0.05% formic acid and acetonitrile mixture as mobile phase with flow rate 1.0 ml/min on gradient mode(Table 2) for the determination of Cabazitaxel and its seven impurities. The optimized chromatographic conditions were shown in Table 3. Cabazitaxelwas eluted at 8.712 ± 0.05 min along with its seven impurities(Figure 2). The individual chromatograms of the impurities (I-VII) were shown in Figure 3.

 

Table. 2. Gradient program of Cabazitaxel

Table. 3. Optimized chromatographic conditions of Cabazitaxel

 

 

 

 

 

 

 

 

 

 

 

 

Figure 2: Characteristic chromatogram of A) Blank B) Cabazitaxel (Rt8.713 min) C) Cabazitaxel (Rt 8.712 min) with its impurities

 

 

Figure 3: Representative chromatograms of Impurities (I-VII) and Blank

Method validation

Cabazitaxelhas shown linearity 0-300 µg/mL (Table 4) with linear regression equation 2813.6x-122.02 (R² = 0.9999) (Figure 4) with % RSD range 0.29-0.94. The linear regression equations of the impurities of Cabazitaxel were shown in Table 5. The % RSD in precision study (n=6) of Cabazitaxel and its impurities was less than 2 confirming that the method is precise (Table 6) and the corresponding chromatograms obtained during the precision study were shown in Figure 5. The percentage recovery of Cabazitaxel and its impurities in accuracy study was shown in Table 7.The % RSD in robustness study of Cabazitaxel and its impurities was less than 2 informing that the method is robust (Table 8) and the corresponding chromatograms obtained during the robustness study were shown in Figure 6.

 

Table. 4. Linearity of Cabazitaxel

*Mean of three replicates

Table. 5.Linearity ofimpurities of Cabazitaxel

 

 

 

Figure 4: Calibration curve of Cabazitaxel

 

 

Table. 6. Precision study of Cabazitaxel and its impurities

Impurities  / Drug	Retention time (min) / Peak area / Theoretical plates (Figure 5)						*Mean peak area ± SD (%RSD)
	1 (A)	2 (B)	3 (C)	4 (D)	5 (E)	6 (F)
I	2.809	2.814	2.81	2.813	2.811	2.803	142964.5 ± 2208.430642 (1.545)
	141126	140791	142830	145739	145603	141698
	15343.576	13882.036	14692.368	13151.873	15298.252	5234.481
II	4.302	4.299	4.294	4.299	4.298	4.294	215336.6667 ± 2967.541384 (1.378)
	211447	213165	214520	215698	219633	217557
	27056.555	26800.334	28406.057	26475.36	26787.453	28664.876
III	5.94	5.93	5.922	5.931	5.933	5.925	105019.1667 ± 1533.300938 (1.889)
	105438	102684	104411	106503	106767	104312
	32151.273	31281.199	30791.995	30531.203	31508.861	32131.726
IV	6.414	6.412	6.407	6.408	6.41	6.404	154465.3333 ± 2666.679708 (1.726)
	155010	151205	158634	153696	155916	152331
	45903.405	47099.344	44881.397	48385.653	48454.409	46381.359
V	7.734	7.733	7.73	7.731	7.733	7.725	183175.1667 ± 3207.670271 (1.751)
	186259	187648	180372	182432	179515	182825
	93411.006	89041.025	97157.477	96519.049	92271.569	91204.197
VI	8.194	8.189	8.184	8.189	8.19	8.18	90076.1667 ± 1400.395432 (1.555)
	90238	90186	90350	92326	89280	88077
	86326.921	83133.742	86489.711	87767.769	85109.1	85968.412
Cabazitaxel	8.732	8.732	8.731	8.732	8.734	8.724	212773.3333 ± 2866.47579 (1.347)
	215967	211862	215525	211774	213339	208173
	123398.05	124401.16	127519.58	129470.47	128868.58	127135.97
VII	10.611	10.614	10.616	10.616	10.617	10.604	144285.5 ± 1110.19093 (0.769)
	143716	142906	143384	145229	145694	144784
	110904.76	103704.84	103926.16	107849	104158.14	107393.18

 

 

 

 

Table. 7. Accuracy study of Cabazitaxel and its impurities

 

 

 

 

 

Table. 8. Robustness study of Cabazitaxel (150 mg/ml)and its impurities (I-VII)

Parameter	Condition	Retention time (min) / Mean peak area / Theoretical plates / (% RSD)
		I	II	III	IV	V	VI	Cabazitaxel	VII
Flow rate (± 0.1 ml/min)	0.8	3.225 183309 12790.899 (0.51)	4.923 231212 28351.943 (0.45)	6.814 109755 34286.024 (0.96)	7.121 190466 61646.476 (0.59)	8.293 198485 98838.779 (0.52)	8.925 102442 89233.545 (0.50)	9.310 264868 123863.642 (0.99)	11.412 172605 100482.543 (0.83)
	1.0	2.822 298059 13936.835 (0.63)	4.314 365564 27174.953 (0.78)	5.951 173713 32463.611 (0.54)	6.427 301875 49010.728 (0.45)	7.743 384054 95454.391 (0.74)	8.192 189155 87034.916 (0.29)	8.742 440391 127218.226 (0.87)	10.625 298288 102318.772 (0.51)
	1.2	2.470 143800 12350.068 (0.55)	3.860 179055 24919.761 (0.69)	5.384 90460 29032.314 (0.65)	5.873 154842 41081.938 (0.82)	7.325 214298 89081.878 (0.65)	7.694 93607 85987.064 (0.86)	8.322 211456 135146.161 (0.96)	10.069 146528 114800.110 (0.39)
Mobile phase ratio Formic acid: acetonitrile (± 5 %, v/v)	Organic phase 5 % less	3.470 174206 29658.879 (0.46)	5.119 217508 36595.893 (0.71)	6.791 102477 47747.904 (0.81)	7.191 180140 78133.198 (0.94)	8.258 186336 127026.549 (0.26)	8.760 99723 106401.682 (0.69)	9.198 265059 139191.920 (0.81)	11.566 178926 85933.619 (0.44)
		2.822 298059 13936.835 (0.39)	4.314 365564 27174.953 (0.38)	5.951 173713 32463.611 (0.69)	6.427 301875 49010.728 (0.87)	7.743 384054 95454.391 (0.89)	8.192 189155 87034.916 (0.75)	8.742 440391 127218.226 (0.57)	10.625 298288 102318.772 (0.54)
	Organic phase 5 % more	2.124 47282 8397.571 (0.45)	3.592 177614 20086.107 (0.85)	5.156 89779 23598.848 (0.72)	5.528 159623 35309.715 (0.39)	7.114 202099 64437.475 (0.67)	7.562 91819 70904.063 (0.53)	8.230 210175 113844.339 (0.39)	9.903 142004 133939.318 (0.55)
Column temperature	20°C	2.812 151332 5513.884 (0.69)	4.294 185225 26572.816 (0.91)	5.922 91970 30025.343 (0.62)	6.406 160169 44788.192 (0.65)	7.731 189545 95699.351 (0.37)	8.185 90775 86939.470 (0.23)	8.733 213183 124171.636 (0.88	10.617 144298 108752.283 (0.25)
	25°C	2.822 298059 13936.835 (0.53)	4.314 365564 27174.953 (0.78)	5.951 173713 32463.611 (0.49)	6.427 301875 49010.728 (0.83)	7.743 384054 95454.391 (0.59)	8.192 189155 87034.916 (0.22)	8.742 440391 127218.226 (0.89)	10.625 298288 102318.772 (0.35)
	30°C	2.807 174480 13991.629 (0.62)	4.291 211963 26784.160 (0.76)	5.917 107593 31225.093 (0.97)	6.404 177507 45246.812 (0.56)	7.727 212852 95939.553 (0.51)	8.150 109164 83299.702 (0.34)	8.727 255749 130084.999 (0.64)	10.613 173535 100468.804 (0.26)

 

Assay of Cabazitaxel

Two different brands of Cabazitaxel formulations were analyzed using both RP-HPLC and LC-ESI-MS methods. Cabazitaxel has shown 99.34-100.05 purity during the assay for the wo brands available in India. The excipients did not interfere with the drug peak.

 

 

0.8 ml/min	1.2 ml/min
Organic phase 5 % less	Organic phase 5 % more
Column temperature at 20°C	Column temperature at 30°C
Figure 6: Characteristic chromatograms of Cabazitaxel and its Impurities (I-VII) during robustness study

 

 

Stress degradation studies of Cabazitaxel

Liquid Chromatographic (RP-HPLC) study

Cabazitaxel and it seven impurities were analyzed using both RP-HPLC and LC-ESI-MS techniques(Total run time of 16 min). Cabazitaxel was eluted at about 8.738 ± 0.05 min in all stress degradation studies. During acidic hydrolysis twodegradant products(DP) were eluted at 2.819 (DP-1) and 9.409 min in the HPLC chromatogram (Figure 7) and during the base hydrolysis study three DPs were observed in the chromatogram at 2.820, 5.932 and 6.429 min. No degradants were marked in oxidation but more than 38% of the drug has undergone degradation andduring photolysis a small DP was observed at 8.978 min. Except oxidation in all the degradation studies less than 15 % of the drug has undergone degradation (Table 9) and the system suitability parameters were within the acceptable criteria. So therefore the mass spectral study was done further to characterize the degradant products.

 

 

Table. 9. Stress degradation studies of Cabazitaxel (RP-HPLC)

Stress conditions	*Mean peak area	Rt (min)	Degradant peaks (DP) (min)		*(%) Drug recovered	* (%) Drug decomposed	Theoretical Plates of drug	Theoretical Plates of DPs
Cabazitaxel	7929456	8.713	-		100	-	129173.970	-
Acidic hydrolysis	7046233	8.738	2.819 9.409		88.86	11.14	118576.979	19952.843 95407.050
Base hydrolysis	7484079	8.735	2.820 5.932 6.429		94.38	5.62	110418.160	19704.216 29939.093 49887.951
Oxidation	4840188	8.734	-		61.04	38.96	120039.686	-
Photolysis	7033047	8.740	8.978		88.69	11.31	115538.741	149954.350
Thermal degradation	7819540	8.737	-		98.61	1.39	113653.970	-
Acid blank				Acid hydrolysis
Base blank				Base hydrolysis
Oxidation blank (H2O2)				Oxidation
Photolysis				Thermal degradation
Figure 7: Characteristic chromatograms of Cabazitaxel during stress degradation studies (RP-HPLC)

 

Liquid Chromatography-Mass spectrophotometric (LC-MS) study

The LC-MS study was further continued to characterize the DPs observed during the degradation studies. All the stress degraded drug mixturesused for the liquid chromatographic study were introduced in to the LC-MS system and the study was continued. The LC-MS chromatogram for the blank solution was shown Figure 8.

 

During the acid hydrolysis the LC-ESI-MS chromatogram (Figure 9)so obtained has shown degradant peaks at 2.83-2.88 (DP-1) and 8.75-8.80 min (Cabazitaxel) and the mass spectra has shown base peaks at m/z values 735.92 and 836.02 respectively(Figure 10).The degradant product DP-1 observed during the acidic hydrolysis has the same retention time as that of Impurity-I eluted at 2.772 min and from the mass spectra it was confirmed that the Impurity I is nothing but the degradant product DP-1. The schematic degradation pathway was shown in Scheme 1 below.

 

During the alkaline hydrolysis the LC-ESI-MS chromatogram (Figure 11) so obtained has shown degradant peaks at 2.886 (DP-1), 5.99 (DP-2), 6.51 min (DP-3) and 8.75-8.88 min (Cabazitaxel) and the mass spectra was shown in Figure 12. The DP2 and DP-3 observed have the same retention time as that of Impurity III and Impurity IV which were confirmed from their mass spectral data. During oxidation and thermal degradation no degradant was seen and therefore mass spectra was not studied.

 

During the photolysis the LC-ESI-MS chromatogram (Figure 13) so obtained has shown degradant peaks at 2.886 (DP-1), 5.99 (DP-2), 6.51 min (DP-3) and 8.75-8.88 min (Cabazitaxel) and the mass spectra was shown in Figure 13. During oxidation and thermal degradation no degradant was seen and therefore mass spectra was not studied.

 

Figure 8: LC-MS Chromatogram of blank

 

Figure 9: LC-ESI-MS chromatogram of Cabazitaxel during acidic hydrolysis

 

 

 

 

Figure 10: LC-ESI-MS chromatogram and Mass spectra of Cabazitaxel during acidic hydrolysis

(Mass spectra of Cabazitaxel (Rt 8.75 min) and DP-1 (Rt 2.83-2.88 min)

 

 

 

Scheme 1: Degradation pathway of Cabazitaxel

 

Figure 11: LC-ESI-MS chromatogram and Mass spectra of Cabazitaxel during base hydrolysis

(Mass spectra of DP-1 at Rt 2.86 min)

 

 

 

Figure 12: LC-ESI-MS chromatogram and Mass spectra of Cabazitaxel during base hydrolysis

(Mass spectra of DP-2 at Rt 5.99 min and DP-3 at Rt 6.51 min)

 

 

 

 

Figure 13: LC-MS chromatogram and Mass spectra of Cabazitaxel during photolysis

(Mass spectra of Cabazitaxel at 8.77 min and DP-4 at Rt 8.98-9.03 min)

 

 

Specificity

Specificity of the method was verified by stress degradation studies. Cabazitaxel peak was not at all interfering with the degradants observed during acidic, alkaline, oxidation, photolysis and thermal degradation studies. The resolution was >2, tailing factor was <1.5 and the theoretical plates were >2000 confirming that the system suitability parameters were satisfied for the HPLC method.

 

CONCLUSIONS:

A new validated stability indicating RP-HPLC method and LC-ESI-MS method were developed for the determination ofCabaitaxeland its seven impurities (I – VII). Three degradant products i.e. DP-1, DP-2 and DP-3 were obtained during the stress degradation studies and they are identical with the Impurities I, III and IV respectively. The mass balance data supported the degradation study and the characterization of the impurities.Theproposed methods werevalidated as per ICH guidelines and found to be sensitiveand specific.

 

ACKNOWLEDGEMENT:

The authors are grateful to Dr. Reddy’s Labs (Hyderabad, India)and Shubham Pharmaceuticalsfor providing the gift samples of Cabazitaxel and its process impurities. The authors have no conflict of interest.

 

REFERENCES:

1.       Oudard S, TROPIC: Phase III trial of Cabazitaxel for the treatment of metastatic castration-resistant prostate cancer, Future Oncology. 7; 2011: 497-506.

2.       Jevtana (cabazitaxel) Injection Approved by U.S. FDA After Priority Review (Press release). sanofi-aventis. 2010-06-17. Retrieved June 17, 2010.

3.       Assessment Report for Jevtana (Cabazitaxel), Procedure No.: EMEA/H/C/002018, European Medicines Agency, London, 2011.

4.       Mathrusri Annapurna M, Pramadvara K, Venkatesh B, Sowjanya G. Stability-indicating RP-HPLC method for the determination of Cabazitaxel. Indo-American Journal of Pharmaceutical Research.  3(11); 2013: 9262-9269.

5.       Mathrusri Annapurna M, Venkatesh B, Supriya GN. A validated stability-indicating liquid chromatographic method for determination of Cabazitaxel - a novel microtubule inhibitor.  Journal of Bioequivalence & Bioavailability.  6(4); 2014: 134-138.

6.       Chengyan Li, Gongjian Lan, Jinyuan Jiang, Mingjie Sun, Taijun Hang. Development and validation of a stability-indicating HPLC method for the determination of the impurities in Cabazitaxel. Chromatographia.  78; 2015: 825-831.

7.       Mathrusri Annapurna M, Venkatesh B, Pramadvara K, Hemchand S. Development and validation of a stability-indicating liquid chromatographic method for the assay of Cabazitaxel. Chemical Science Transactions. 3(2); 2014:854-860.

8.       Venkatesh B, Mathrusri Annapurna M, Pramadvara K. Analytical stress degradation studies of Cabazitaxel (a semi synthetic natural taxoid) using liquid chromatography. Pharmaceutical Methods. 6(3); 2015:137-142.

9.       Mathrusri Annapurna Mukthinuthalapati, Venkatesh Bukkapatnam, Sunkara Mrunal Chaitanya. Determination of Cabazitaxel (an anti-prostate cancer agent) by reverse phase liquid chromatography. Research Journal of Pharmacy and Technology. 10(4); 2017: 1138-1144.

10.     Mathrusri Annapurna Mukthinuthalapati, Venkatesh Bukkapatnam, A new validated RP-HPLC method for the quantification of Cabazitaxel - an anticancer plant product. Science Spectrum.  2(1); 2017: 31-38.

11.     Kort A, Hillebrand MJX, Cirkel GA, Voest EE, Schinkel AH, Rosing H, Schellen JHM, Beijnen JH. Quantification of Cabazitaxel, its metabolite docetaxel and the determination of the demethylated metabolites RPR112698 and RPR123142 as docetaxel equivalents in human plasma by liquid chromatography tandem mass spectrometry. Journal of Chromatography B. 925; 2013:117-123.

12.     Peter de Bruijn, Anne-Joy M de Graan, AnnemiekeNieuweboer, Ron HJ Mathijssen, Mei-Ho Lam, Ronald de Wit, Erik, AC Wiemer, Walter J Loos. Quantification of Cabazitaxel in human plasma by liquid chromatography/ triple-quadrupole mass spectrometry: a practical solution for non-specific binding. Journal of Pharmaceutical and Biomedical Analysis. 59; 2012: 117–122.

13.     Jagannath Patro1 V, Nageshwara Rao R, Tripathy NK.  LC-MS/MS determination of Cabazitaxel in rat whole blood on dry blood Spots. Open Access Scientific Reports. 1; 2012: 1-4.

14.     ICH Q2 (R1) Validation of analytical procedures: Text and methodology (2005).

15.     ICH Q1A (R2) Stability testing of new drug substances and products (2003).

16.     ICH Q3A (R2) Impurities in new drug substances (2006).

 

 

 

 

 

Accepted on 04.10.2018        © RJPT All right reserved

Research J. Pharm. and Tech 2018; 11(10): 4683-4698.