UF PHA6936 FINAL EXAM PREP TEST
BANK ACTUAL QUESTIONS AND VERIFIED
RESPONSES FULL SOLUTION
⩥ Mass Spectrometry
Answer: the sample is in vapor form and is bombarded with e-, which
converts the substance to an ionic state which consists of the parent ion
and several ionic fragments of the original molecule. They are separated
according to their mass/charge ratio. The MW of the compound can be
deter from it's spectrum.
ADV. --specific- best analytical technique, sensitive ng/ml, qual and
quant, can be used with separation techniques
DIS--expensive, high maitenence, trained analysts, less sensitive than
Fluorescence, best for low MW < 500, can be issues LC
⩥ Validation of Analytical Procedures
Answer: key parameters in the validation process are:
1. purity and identity of the neat test sample
2. homogeneity of soltns
3. extraction recovery
4. range of standard curve and control samples
,5. linearity and precision of standard curves
6. sensitivity
7. specificity
8. accuracy and precision
9. stability
10. quality control
⩥ Validation of Analytical Procedures: ID and purity of test compound
Answer: PURITY --this is usually done via GC or HPLC and the
analysis of the sample should show the presence of a single major peak
with a retention time corresponding to that of previously injected
standards
IDENTITY--done via MS
⩥ Validation of Analytical Procedures: homogeneity of soltns
Answer: taking aliquots from the top, middle and bottom of a sample
ctgr. Concentrations should fall in an acceptable range.
⩥ Validation of Analytical Procedures: Extraction Recovery
Answer: this is deter by the comparison of the peak area observed with a
soltn prepared at the same conc in the appropriate matrix, it is then
extracted and injected into the chromatographic system. Three
concentrations should be used.
,⩥ Validation of Analytical Procedures: Range of Std. Curve and Control
Samples
Answer: sufficient vol of the std curve and control samples should be
prepared in a drug free pooled serum (or other bio matrix) for use of the
remainder of the study. Std curve calibrators are prepared for each
compound of interest in each biological matrix to be analzyed during the
study.
⩥ Validation of Analytical Procedures: Sensitivity
Answer: this is measured by deter the limit of detection (LOD). Can be
done several ways such as: analyzing samples of known conc and est the
min level at which the analyte can be reliably detected. OR using signal
to noise ratios by comparing measured signals from samples with known
low concentrations of analyte with those of blank samples and est the
minimum conc at which the analyte can be detected, blanks must be
analyzed to est background noise
⩥ Validation of Analytical Procedures: Quantitation Limit
Answer: this is done by the analysis of samples with known
concentrations of analyte and est the min level at which the analyte can
be quantified with acceptable accuracy and precision. A typical signal to
noise ratio is 10:1
⩥ Validation of Analytical Procedures: Specificity
, Answer: this is when a test can discriminate bt compounds of closely
related structures like drug metabolites. This can be demonstrated by
obtaining positive results from the samples ctg the analyte and negative
results from the samples that do not ctg the analyte. The assay should be
applied to a like substance to check for specificity.
⩥ Validation of Analytical Procedures: Accuracy and Precision
Answer: Accuracy-- can be verified by comparison of the results of the
proposed analytical procedure with those of a second well-characterized
procedure. should be done using at least 9 determinations with three
conc levels. Generally reported as the diff bt the mean and the accepted
true value together with the confidence intervals.
Precision--replicate analysis of 3 with varied conc. control samples are
performed on the same day over a minimum of two weeks
⩥ Validation of Analytical Procedures: Stability
Answer: parameters that should be heeded should be included in the
report such as pH, temp, humidity. Stability of a standard curve, control
and subject samples is prospectively demonstrated for the sample
collection and storage conditions used for the study
biological samples need to be validated with replicate injections of the
sample over 24 hours
⩥ Validation of Analytical Procedures: Quality Control
BANK ACTUAL QUESTIONS AND VERIFIED
RESPONSES FULL SOLUTION
⩥ Mass Spectrometry
Answer: the sample is in vapor form and is bombarded with e-, which
converts the substance to an ionic state which consists of the parent ion
and several ionic fragments of the original molecule. They are separated
according to their mass/charge ratio. The MW of the compound can be
deter from it's spectrum.
ADV. --specific- best analytical technique, sensitive ng/ml, qual and
quant, can be used with separation techniques
DIS--expensive, high maitenence, trained analysts, less sensitive than
Fluorescence, best for low MW < 500, can be issues LC
⩥ Validation of Analytical Procedures
Answer: key parameters in the validation process are:
1. purity and identity of the neat test sample
2. homogeneity of soltns
3. extraction recovery
4. range of standard curve and control samples
,5. linearity and precision of standard curves
6. sensitivity
7. specificity
8. accuracy and precision
9. stability
10. quality control
⩥ Validation of Analytical Procedures: ID and purity of test compound
Answer: PURITY --this is usually done via GC or HPLC and the
analysis of the sample should show the presence of a single major peak
with a retention time corresponding to that of previously injected
standards
IDENTITY--done via MS
⩥ Validation of Analytical Procedures: homogeneity of soltns
Answer: taking aliquots from the top, middle and bottom of a sample
ctgr. Concentrations should fall in an acceptable range.
⩥ Validation of Analytical Procedures: Extraction Recovery
Answer: this is deter by the comparison of the peak area observed with a
soltn prepared at the same conc in the appropriate matrix, it is then
extracted and injected into the chromatographic system. Three
concentrations should be used.
,⩥ Validation of Analytical Procedures: Range of Std. Curve and Control
Samples
Answer: sufficient vol of the std curve and control samples should be
prepared in a drug free pooled serum (or other bio matrix) for use of the
remainder of the study. Std curve calibrators are prepared for each
compound of interest in each biological matrix to be analzyed during the
study.
⩥ Validation of Analytical Procedures: Sensitivity
Answer: this is measured by deter the limit of detection (LOD). Can be
done several ways such as: analyzing samples of known conc and est the
min level at which the analyte can be reliably detected. OR using signal
to noise ratios by comparing measured signals from samples with known
low concentrations of analyte with those of blank samples and est the
minimum conc at which the analyte can be detected, blanks must be
analyzed to est background noise
⩥ Validation of Analytical Procedures: Quantitation Limit
Answer: this is done by the analysis of samples with known
concentrations of analyte and est the min level at which the analyte can
be quantified with acceptable accuracy and precision. A typical signal to
noise ratio is 10:1
⩥ Validation of Analytical Procedures: Specificity
, Answer: this is when a test can discriminate bt compounds of closely
related structures like drug metabolites. This can be demonstrated by
obtaining positive results from the samples ctg the analyte and negative
results from the samples that do not ctg the analyte. The assay should be
applied to a like substance to check for specificity.
⩥ Validation of Analytical Procedures: Accuracy and Precision
Answer: Accuracy-- can be verified by comparison of the results of the
proposed analytical procedure with those of a second well-characterized
procedure. should be done using at least 9 determinations with three
conc levels. Generally reported as the diff bt the mean and the accepted
true value together with the confidence intervals.
Precision--replicate analysis of 3 with varied conc. control samples are
performed on the same day over a minimum of two weeks
⩥ Validation of Analytical Procedures: Stability
Answer: parameters that should be heeded should be included in the
report such as pH, temp, humidity. Stability of a standard curve, control
and subject samples is prospectively demonstrated for the sample
collection and storage conditions used for the study
biological samples need to be validated with replicate injections of the
sample over 24 hours
⩥ Validation of Analytical Procedures: Quality Control
