Stability Indicating Assays for Dose Formulation Stability Testing Teresa Fuller GSK.
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Transcript of Stability Indicating Assays for Dose Formulation Stability Testing Teresa Fuller GSK.
Stability Indicating Assays for Dose Formulation Stability Testing
Teresa FullerGSK
Stability Indicating Assays for Dose Formulation Stability Testing
– Stability Studies for Dose Formulations
– What the white paper says
– What is a stability indicating assay?
– So do we need a stability-indicating assay?
– Closing thoughts
Stability Studies for Formulations
– Performed for every formulation to be used on a GLP study
– Test the formulation for concentration on day 0
– Test the formulation for concentration on further timepoints to assess stability, eg. day 1, day 7 and day 14
– Based on concentration results over time, assess stability
What the white paper says
“The analytical method should be suitable for the intended purpose. It is conceivable to have a rapid analytical method for test article only, which would not be suitable as a stability-indicating assay where degradents are separated from the test article or a more elaborate method which may have the additional advantage of identifying degradents. Both types of assays are acceptable and would require the appropriate level of validation for their intended purposes.”
What is a Stability Indicating Assay?
– According to the US-FDA stability guideline of 1998
‘validated quantitative analytical methods that can detect the changes with time in the chemical, physical, or microbiological properties of the drug substance and drug product, and that are specific so that the contents of active ingredient, degradation products, and other components of interest can be accurately measured without interference.’
– But this applies to drug substance and drug product not dose formulations!
– For dose formulations we do not necessarily need to quantify the degradents
– So what should the formulation analytical method look like?
Isocratic UPLC/UV
– Chromatography column, %organic solvent and wavelength chosen for analyte of interest
– K’ of at least 3 [K’ = (Vt – V0)/V0]
– Eg.
HPLC System Waters Acquity UPLC with TUV Detector using High Sensitivity Flow cell
Typical Injection Volume 1 µL
Flow Rate 1 mL/min
Wavelength 245 nm
Analytical Column 50 x 2.1 mm i.d. Waters BEH C18 1.7 µm
Column Temperature 60 ºC
Run Time 1 min
Typical Retention Time 0.8 min
Mobile Phase A 0.05% Trifluoroacetic Acid
Mobile Phase B Methanol
Isocratic Composition A:B 69:31
Isocratic UPLC/UV
- 0
.4
9
- 0
.7
0
GS
K3
00
83
48
- 0
.7
64.0
5.0
6.0
7.0
8.0
9.0
10.0
11.0
0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35 0.40 0.45 0.50 0.55 0.60 0.65 0.70 0.75 0.80 0.85 0.90 0.95
GS
K3008348
- 0.7
5
4.0
5.0
6.0
7.0
8.0
9.0
10.0
0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35 0.40 0.45 0.50 0.55 0.60 0.65 0.70 0.75 0.80 0.85 0.90 0.95
Reference standard
Stability sample day 7, ambient PFL0.02 mg/mL in water
Generic UPLC-UV Gradient
HPLC System Waters Acquity UPLC
Typical Injection Volume 0.5 – 5 uL
Flow Rate 0.6 mL/min
Wavelength Analyte specific
Analytical Column 100 x 2.1 mm i.d. Waters BEH C18 1.7 µm
Column Temperature 50 ºC
Run Time 12 min
Mobile Phase A 0.1% Trifluoroacetic Acid in water
Mobile Phase B 0.1% Trifluoroacetic Acid in acetonitrile
Gradient profile Time %B
0 3
10 100
10.5 100
11 3
Generic UPLC-UV Gradient
Isocratic and Gradient Compared
Isocratic Gradient
Run Time 1-2 mins 12 mins
Method development 1 day to several weeks Minimal
Visible degradents Significantly different degradents probably not visible
Most degradents and impurities likely to be visible
Stability-indicating possibly possibly
How can we assess how stability indicating the assay is?
– Forced degradation
– Store sample at 30°C, 65% RH or 40°C, 75% RH or 50°C, ambient RH
– UV light exposure
– Oxidation, eg. By addition of hydrogen peroxide
– Acid/alkali hydrolysis
– Compare forced samples to a fresh sample
– If any additional peaks are seen in the forced samples but these are well resolved from the analyte then assume assay is adequate
How can we assess how stability indicating the assay is?
– Peak purity analysis
– Acquire UV spectrum at several points (at least 3) across the chromatographic peak
– Compare spectra to look for differences which may indicate underlying peaks co-eluting with the analyte
How can we assess how stability indicating the assay is?
– Mass spectral peak purity
– Acquire mass spectrum at several points across the peak, similarly to UV peak purity
The ideal stability indicating method?
– UPLC-MS/MS
– Use specific parent and daughter ions to monitor the analyte
Analyte Precursor ion(m/z)
Product Ion (m/z)
GSK123456 539 290
Product 290
XIC of +MRM (2 pairs): 391.0/154.2 amu from Sample 3 (STD5) of GSK2245035KMSLIVALB-... Max. 1.2e4 cps.
0.5 1.0 1.5Time, min
0.00
2000.00
4000.00
6000.00
8000.00
1.00e4
1.20e4
Inte
ns
ity, c
ps
1.14
Pros and Cons
Technique Pros Cons
Forced Degradation
•No additional equipment reqd
•Time•Degradation may not be the same in real samples
UV Peak purity •Quick •Degradation products may have similar spectrum•Needs PDA detector
Mass spec •Specific •Expensive equipment•Mass spectral comparison difficult
So do we need a stability-indicating assay?
– White paper says we don’t!
– Unstable formulation considered to be greater than 10% drop from t0
– Complex oral and iv formulations are likely to have had previous development performed by a Product Development group
– storage stability already tested to some degree
– We need some confidence in our assay
Closing Thoughts
– Several factors to consider
– Balancing act between having confidence in the method and minimising method development time and effort
– Is a gradient better than isocratic?
– Is MS detection better than UV as it becomes more affordable?
Acknowledgements
– Emma Simmonds, GSK PD
– Tammy Clegg, GSK DMPK
– Louise Kay, GSK DMPK
Useful Reference
– Review of Development of Validated Stability Indicating Assay Methods
– Journal of Pharmaceutical and Biomedical Analysis 28 (2002) 1011–1040