Table 1.
Analytical Method Validationa and Stability Data for Vinpocetine and Apovincaminic Acid in Dam Plasma, Amniotic Fluid, and Fetal Homogenate
| Plasma | Amniotic Fluid | Fetal Homogenate | |
|---|---|---|---|
| Vinpocetine | |||
| Concentration Range (ng/mL or ng/g)b | 0.5 to 100 | 0.5 to 100 | 5-1000 |
| Linearity (r)c | ≥ 0.999 | 0.999 | 0.998 |
| LOQ (ng/mL or ng/g)d,e | 0.5 | 0.5 | 5 |
| LOD (ng/mL or ng/g)f | 0.05 | 0.05 | 0.7 |
| Accuracy (%RE) | |||
| Intra-day | −3.3 to 4.3 | −7.5 to 12.7 | −7.3 to 5.1 |
| Inter-day | −2.3 to 2.3 | NA | −2.0 to 2.7 |
| Precision (%RSD) | |||
| Intra-day | 0.7 to 9.6 | 0 to 4.7 | 0.5 to 8.9 |
| Inter-day | 1.7 to 6.1 | NA | 1.4 to 8.3 |
| Absolute Recovery (%)g | 89.6 to 124.4 | 81.2 to 102.0 | 103.8 to 150.3 |
| Dilution Verification, n=6 (% RE, %RSD)h | −7.9, 5.9 | 4.0, 2.3 | 6.9, 7.6 |
| Extracted Sample Stability, n=4 (%RE)i | |||
| Ambient temperature (7d)j | −7.5 to −12.9 | −0.2 to 12.4 | −8.8 to −0.4 |
| Freeze-thaw (3 cycles) | −16.5 to −6.0 | 0.4 to 8.6 | −8.8 to −2.9 |
| Matrix Stability (%RE)k | |||
| Freeze-Thaw (3 cycles) | −14.8 to −14.7 | −8.7 to −3.0 | 1.3 to 4.7 |
| Storage (61d) | −12.9 to −8.4 | −13.9 to −9.1 | 4.2 to 6.0 |
| Apovincaminic Acid | |||
| Concentration Range (ng/mL or ng/g)b | 0.5 to 100 | 0.5 to 100 | |
| Linearity (r)c | ≥ 0.995 | 0.997 | 0.998 |
| LOQ (ng/mL or ng/g)d,e | 0.5 | 0.5 | 5 |
| LOD (ng/mL or ng/g)f | 0.0567 | 0.140 | 0.777 |
| Accuracy (%RE) | |||
| Intra-day | −16.3 to 16.3 | −7.6 to 7.8 | −2.8 to 6.9 |
| Inter-day | −7.8 to 6.7 | NA | −1.6 to 2.0 |
| Precision (%RSD) | |||
| Intra-day | 0 to 23.7 | 0-10.7 | 0.8 to 15.2 |
| Inter-day | 3.2 to 14.4 | NA | 2.8 to 9.8 |
| Absolute Recovery (%)g | 160.2 to 249.7 | 131 to 172 | 119.3 to 159.3 |
| Dilution Verification, n=6 (% RE, %RSD)h | 3.9, 8.9 | −4.9, 8.5 | −2.9, 3.7 |
| Extracted Sample Stability, n=4 (%RE) i | |||
| Ambient temperature (7d)j | −2.4 to −5.4 | −7.3 to 14.7 | −14.1 to −2.4 |
| Freeze-thaw (3 cycles) | −13.7 to 3.6 | −10.5 to 5.7 | −9.8 to 2.7 |
| Matrix Stability (% RE)k | |||
| Freeze-Thaw (3 cycles) | −7.6 to 10.6 | 5.2 to 10.2 | −9.1 to 2.2 |
| Storage (61d) | −22.0 to −12.4 | −11.4 to −1.9 | −7.5 to −2.2 |
Full validations were performed in plasma and fetal homogenate. A partial validation was performed for amniotic fluid using plasma method.
Plasma and amniotic fluid concentrations are expressed as ng/mL and fetal homogenate as ng/g.
For both vinpocetine and AVA, plasma and amniotic fluid curves were fitted with linear 1/x weighted regression and fetal homogenate curves were fitted with linear 1/x2 weighted regression.
LOQ, limit of quantitation; LOD, limit of detection, RE, relative error; RSD, relative standard deviation; NA, not applicable.
Experimental LOQ, is the lowest concentration used in standard curve. Target values are given.
Estimated as the 3 times the standard deviation of the LOQ (n=6 replicates).
Values given are the range for the concentration range validated.
Highest concentration verified were: plasma and amniotic fluid, 1000 ng/mL; fetal homogenate, 10,000 ng/g
Values given are the range for 3 QC concentrations at 1, 30 and 75 ng/mL in plasma and amniotic fluid and 7, 320 and 750 ng/g in fetal homogenate.
Values given are mean percent recovered after refrigerator, autosampler or ambient temperature storage at least 7 days
Values given are for 2 target QC concentrations at 1 and 75 ng/mL in plasma and amniotic fluid and 7 and 750 ng/g in fetal homogenate.