High-throughput method to analyze tegafur and 5-fluorouracil in human tears and plasma using hydrophilic interaction liquid chromatography/tandem mass spectrometry.

RATIONALE: We developed a new high-throughput method to analyze tegafur (FT) and 5-fluorouracil (5-FU) in tear and plasma samples using hydrophilic interaction liquid chromatography (HILIC)/tandem mass spectrometry (MS/MS). METHODS: The tear samples (10 µL) spiked with FT, 5-FU, and 5-chlorouracil (internal standard) were diluted using 40 µL of 2 M ammonium acetate and 250 µL of acetonitrile with 2% formic acid; 20 µL of plasma spiked with the two drugs and internal standard was diluted with 80 µL of 2 M ammonium acetate and 500 µL of acetonitrile with 2% formic acid. After centrifugation, the clear supernatant extract (15 µL) was directly injected into the HILIC/MS/MS instrument, and each drug was separated on a Unison UK-Amino column (50 mm × 3 mm i.d., 3 µm particle size) with a linear gradient elution system composed of 10 mM ammonium acetate (pH 6.8) and acetonitrile at a flow rate of 0.7 mL/min. We performed quantification by multiple reaction monitoring (MRM) with negative-ion atmospheric-pressure chemical ionization. RESULTS: Distinct peaks were observed for the drugs on each MRM channel within 2 min. The regression equations showed good linearity within the range 0.04-4.0 µg/mL for the tear and plasma samples with detection limits at 0.02-0.04 µg/mL. Recoveries for target analytes (FT and 5-FU) for the tear and plasma samples were in the 94-128% and 94-104% ranges, respectively. The intra- and inter-day coefficients of variation for the two drugs were lower than 10.8%. The accuracies of quantitation were 97-115% for both samples. CONCLUSIONS: We established a high-throughput, reproducible, and practical procedure for analyzing FT and 5-FU in human tear and plasma samples using HILIC/MS/MS analysis with an aminopropyl-bonded mixed-mode separation column. This method can be applied to the high-throughput routines used in clinical analyses.

Volume of the right supramarginal gyrus is associated with a maintenance of emotion recognition ability.

Emotion recognition is known to change with age, but associations between the change and brain atrophy are not well understood. In the current study atrophied brain regions associated with emotion recognition were investigated in elderly and younger participants. Group comparison showed no difference in emotion recognition score, while the score was associated with years of education, not age. We measured the gray matter volume of 18 regions of interest including the bilateral precuneus, supramarginal gyrus, orbital gyrus, straight gyrus, superior temporal sulcus, inferior frontal gyrus, insular cortex, amygdala, and hippocampus, which have been associated with social function and emotion recognition. Brain reductions were observed in elderly group except left inferior frontal gyrus, left straight gyrus, right orbital gyrus, right inferior frontal gyrus, and right supramarginal gyrus. Path analysis was performed using the following variables: age, years of education, emotion recognition score, and the 5 regions that were not different between the groups. The analysis revealed that years of education were associated with volumes of the right orbital gyrus, right inferior frontal gyrus, and right supramarginal gyrus. Furthermore, the right supramarginal gyrus volume was associated with the emotion recognition score. These results suggest that the amount of education received contributes to maintain the right supramarginal gyrus volume, and indirectly affects emotion recognition ability.

Relationship between vascular endothelial growth factor and macular edema in retinal vein branch obstruction.

PURPOSE: Vascular endothelial growth factor (VEGF) plays an important role in branch retinal vein occlusion (BRVO) with cystoid macular edema (CME). Monitoring changes in VEGF is crucial for evaluating treatment but requires vitreous or aqueous humor sampling, which hampers its clinical application. We investigated the correlation between VEGF and protein concentration in the aqueous humor (flare) and whether this could be used to monitor treatment-related VEGF changes. DESIGN: This retrospective observational study involved 19 previously untreated patients with BRVO. Aqueous humor was obtained, and intravitreal ranibizumab (IVR) injection was administered to these patients. The correlation between VEGF and flare, central retinal thickness (CRT), and best-corrected visual acuity (BCVA) was investigated. Differences in these values were considered between pre-IVR and 1 week and 1-3 months post-IVR. Moreover, in patients with recurrence who received additional IVR, further changes in VEGF were examined. MAIN OUTCOME MEASURES: The end point of this study was BCVA, flare, and CRT at the fovea. RESULTS: Significant improvement was seen in BCVA and CRT at all time points and in Flare at 1 vs 3 months post-IVR; nevertheless, additional IVR was necessary in 94% of cases. In a patient with recurrence, CRT did not improve, even though VEGF decreased. CONCLUSION: Flare may be effective for estimating VEGF levels in aqueous humor pre-IVR. Inflammation-related molecules other than VEGF may be related to recurrence.

Effects of green tea fractions on oxygen-induced retinal neovascularization in the neonatal rat.

This study aimed to investigate the preventive effects of green tea fractions (GTFs) on rat model of oxygen-induced retinopathy (OIR). Neonatal Sprague-Dawley rats were exposed to daily cycles of 80% oxygen (20.5 h), ambient air (0.5 h), and progressive return to 80% oxygen (3 h) until postnatal day 12 (P12), then the rats were placed in ambient air until P18. The green tea was fractionated by DM-A50, DM-W, M-B, and M-W. The rats were treated once daily from P6 to P17 by gastric gavage of GTFs (0.05 or 0.01 g/ml) or distilled water (DW) at 50 microl/10 g body weight. On P18, the rats were sacrificed and the retinal samples were collected. The retinal neovascularization (NV) was scored and avascular areas (AVAs) were measured as a % of total retinal area (%AVAs) in ADPase stained retinas. The NV scores in 0.01 g/ml M-W were significantly lower than those in DW. The %AVAs in 0.05 g/ml DM-A50 and in 0.05 g/ml and 0.01 g/ml M-W were significantly lower than those in DW. There were less catechins, and less caffeine in M-W fraction compared with other GTFs, suggesting components of green tea except for catechins and caffeine might suppress the neovascularization in rat model of OIR.

[The effect of Sairei-to on oxygen-induced retinal neovascularization in the neonatal rat].

PURPOSE: To study the effect of Sairei-to (ST), a Japanese traditional medicine, on oxygen-induced retinopathy (OIR) in rats. METHODS: OIR was induced by maintaining Sprague-Dawley neonatal rats in 80% oxygen for 12 days. The rats were treated once daily with oral administration of 0.75 g/kg (n = 9), 1.5 g/kg (n = 13) of ST in water, or water alone (WA, n = 13) at 5 mL/ kg body weight from day 6 to day 17. On day 18, retinal samples were collected. Retinal neovascularization (NV) was assessed by the NV score, and by the percentage of avascular area (% AVA), using a method previously reported. The number of severe retinal NV cases (NV > or = 9) was compared. The retinal vascular endothelial growth factor (VEGF) concentrations were measured with an immunoassay kit, at 0, 12, 24, 72 and 144 hours after oxygenation. RESULTS: NV score and % AVA decreased in the ST treated group compared to the WA group. However, severe NV was seen in five cases of WA and in one case of the ST treated group. Thus severe NV was inhibited significantly by ST treatment (p = 0.0185). Retinal VEGF did not differ between groups at any time points. CONCLUSION: These data suggest that severe NV in OIR is inhibited by ST treatment.