In the current work, a simple, economical, accurate, and precise HPLC method with UV detection was developed to quantify Favipiravir (FVIR) in spiked human plasma using acyclovir (ACVR) as an internal standard in the COVID-19 pandemic time. Both FVIR and ACVR were well separated and resolved on the C18 column using the mobile phase blend of methanol:acetonitrile:20 mM phosphate buffer (pH 3.1) in an isocratic mode flow rate of 1 mL/min with a proportion of 30:10:60 %, v/v/v. The detector wavelength was set at 242 nm. Maximum recovery of FVIR and ACVR from plasma was obtained with dichloromethane (DCM) as extracting solvent. The calibration curve was found to be linear in the range of 3.1–60.0 µg/mL with regression coefficient (r²) = 0.9976. However, with acceptable r², the calibration data’s heteroscedasticity was observed, which was further reduced using weighted linear regression with weighting factor 1/x. Finally, the method was validated concerning sensitivity, accuracy (Inter and Intraday’s % RE and RSD were 0.28, 0.65 and 1.00, 0.12 respectively), precision, recovery (89.99%, 89.09%, and 90.81% for LQC, MQC, and HQC, respectively), stability (% RSD for 30-day were 3.04 and 1.71 for LQC and HQC, respectively at −20 °C), and carry-over US-FDA guidance for Bioanalytical Method Validation for researchers in the COVID-19 pandemic crisis. Furthermore, there was no significant difference for selectivity when evaluated at LLOQ concentration of 3 µg/mL of FVIR and relative to the blank.

The aim of this study was to evaluate the relationship between the protamine ratio (P1/P2), DNA fragmentation of spermatozoa and protamine deficiency. Patients were grouped into fertile (G1; n = 151) and sub‐fertile (G2; n = 121). DNA fragmentation in spermatozoa was analysed by a TUNEL assay (terminal deoxynucleotidyl transferase‐mediated deoxyuridine triphosphate nick‐end labelling), and the protamination was determined by CMA3 staining, while Western blot was used to measure protamine P1 and P2. While sperm DNA fragmentation (SDF) and protamine ratio were significantly elevated in G2 compared with G1 (12.31 ± 7.01% vs. 17.5 ± 9.5%; p = .001) and (0.91 ± 0.43 vs. 0.75 ± 0.42; p = .003); respectively, the CMA3 positive showed no difference at all between G1 and G2. In G1, the CMA3 positive correlated negatively with the P1/P2 ratio and SDF (r = −.586, r = −.297; p = .001 respectively). In contrast, the protamine ratio correlated positively with SDF (r = .356; p = .001). In G2, no correlation was observed between CMA3 positive, SDF and the P1/P2 ratio but the P1/P2 ratio showed a positive correlation with SDF (r = .479; p = .001). In conclusion, the spermatozoa DNA deterioration was closely associated with abnormal protamination but showed an association with the protamine ratio, more than with CMA3 positive. Therefore, for the evaluation of DNA damage in spermatozoa, the P1/P2 ratio might act as an additional biomarker.

A novel, sensitive, and low-cost HPLC method for the rapid determination of favipiravir (FVR) in rat plasma was developed and validated, and the effect of vitamin C on FVR pharmacokinetic parameters was investigated. FVR and oxcarbazepine (IS) were separated using a mobile phase of 50% acetonitrile and 50% water (with 0.25% trifluoroacetic acid) at 1.0 mL/min flow rate and detected at λmax 289 nm. The intra- and interday values for FVR in plasma were less than 15%, with low, medium, and high QC levels for the relative recovery rate, according to ICH guidelines. Cmax values in the control and experimental groups were 558 ± 124.42 and 979.13 ± 138.10 ng/mL, respectively; t_1/2 values were 7.15 ± 1.60 and 9.09 ± 1.14 h, AUC_(0-t) values were 5697.70 ± 536.58 and 7381.62 ± 1577.58 ng.h/mL, and AUC_(0-∞) values were 5697.70 ± 536.58 and 8192.36 ± 1721.67, respectively. According to the results, the experimental group’s Cmax of FVR was 75.17% higher than the control group’s, the Vz/F was lower, and the t_1/2 was 1.86 h longer. The technique developed for determining FVR in plasma was useful for FVR pharmacokinetics and food–drug interaction investigations.