Table 5.
Assessment of effects of deionized water on the extraction efficiency of eicosanoids and on the reproducibility of the assay. Data are represented as the average extraction recovery (E.R) values for deionized water. Similarly matrix effect (M.E) of deionized water on eicosanoids was also measured. Data are represented as the average IS-normalized ME values and the CV for deionized water spiked with low and high concentrations of each eicosanoid at 0.417 ng/ml (25 pg on column) and 8.33 ng/ml (500 pg on column). IS-normalized M.E values in deionized water were compared to buffer. Results indicate overall improvement in extraction recovery of eicosanoids and reduction in matrix effects when deionized water was used as compared to microsomal incubation buffer.
| Medium | Analyte | Amt (pg) | M.E | E.R | ||
|---|---|---|---|---|---|---|
|
| ||||||
| Mean | %C.V | Mean | %C.V | |||
| Buffer | 20-HETE | 25 | 0.94 | 11.2 | 113.5 | 18.7 |
| 12-HETE | 25 | 0.80 | 12.2 | 108.4 | 10.1 | |
| 15-HETE | 25 | 0.93 | 8.3 | 85.2 | 12.8 | |
| (n=5) | 8,9-EET | 25 | 1.09 | 13.4 | 98.3 | 16.2 |
| 11,12-EET | 25 | 0.92 | 13.8 | 104.2 | 11.6 | |
| 14,15-EET | 25 | 0.91 | 8.5 | 111.4 | 13.7 | |
| 5,6-DiHETE | 25 | 1.03 | 2.4 | 104.0 | 17.6 | |
| 8,9-DiHETE | 25 | 1.01 | 5.9 | 107.0 | 12.6 | |
| 11,12-DiHETE | 25 | 0.97 | 7.3 | 117.0 | 5.2 | |
| 14,15-DiHETE | 25 | 1.11 | 4.8 | 115.7 | 5.9 | |
| Water | 20-HETE | 25 | 1.08 | 5.6 | 104.6 | 10.7 |
| 12-HETE | 25 | 1.06 | 15.6 | 95.8 | 14.7 | |
| 15-HETE | 25 | 1.00 | 8.4 | 92.4 | 10.4 | |
| (n=5) | 8,9-EET | 25 | 1.04 | 10.8 | 104.3 | 10.0 |
| 11,12-EET | 25 | 1.03 | 16.9 | 103.3 | 15.0 | |
| 14,15-EET | 25 | 0.99 | 10.8 | 98.7 | 18.2 | |
| 5,6-DiHETE | 25 | 1.02 | 4.5 | 111.2 | 19.7 | |
| 8,9-DiHETE | 25 | 1.17 | 6.3 | 92.0 | 12.5 | |
| 11,12-DiHETE | 25 | 1.01 | 6.6 | 109.5 | 9.9 | |
| 14,15-DiHETE | 25 | 1.06 | 8.9 | 105.4 | 17.0 | |
| Buffer | 20-HETE | 500 | 1.04 | 6.0 | 99.2 | 10.6 |
| 12-HETE | 500 | 0.90 | 14.4 | 103.5 | 10.3 | |
| 15-HETE | 500 | 0.93 | 11.7 | 97.6 | 9.8 | |
| 8,9-EET | 500 | 0.97 | 16.0 | 97.1 | 12.2 | |
| (n=5) | 11,12-EET | 500 | 0.95 | 15.8 | 101.5 | 9.1 |
| 14,15-EET | 500 | 0.97 | 14.8 | 99.8 | 12.3 | |
| 5,6-DiHETE | 500 | 0.99 | 15.3 | 105.3 | 12.6 | |
| 8,9-DiHETE | 500 | 0.98 | 10.7 | 106.2 | 9.5 | |
| 11,12-DiHETE | 500 | 1.02 | 12.4 | 106.9 | 12.0 | |
| 14,15-DiHETE | 500 | 1.07 | 7.7 | 108.3 | 6.6 | |
| Water | 20-HETE | 500 | 0.95 | 7.8 | 103.2 | 5.0 |
| 12-HETE | 500 | 1.06 | 10.8 | 98.1 | 7.0 | |
| 15-HETE | 500 | 1.03 | 7.5 | 98.9 | 7.8 | |
| (n=5) | 8,9-EET | 500 | 1.05 | 6.8 | 101.1 | 6.6 |
| 11,12-EET | 500 | 1.07 | 11.2 | 101.6 | 3.5 | |
| 14,15-EET | 500 | 1.03 | 7.6 | 99.4 | 5.2 | |
| 5,6-DiHETE | 500 | 1.02 | 1.1 | 111.0 | 5.6 | |
| 8,9-DiHETE | 500 | 1.03 | 8.1 | 112.1 | 4.5 | |
| 11,12-DiHETE | 500 | 1.06 | 7.9 | 104.6 | 6.3 | |
| 14,15-DiHETE | 500 | 1.07 | 1.9 | 105.5 | 3.3 | |