In vitro Inhibition Kinetics of Crocodylus mindorensis (Philippine crocodile) Serum against Human Immunodeficiency Virus Type I Reverse Transcriptase

Abstract

The study aimed to determine the inhibition kinetics of Crocodylus mindorensis (Philippine crocodile) serum against recombinant HIV-1 Reverse Transcriptase (RT) using Enzyme-linked Immunosorbent Assay. The study utilized an experimental design where different concentrations of Philippine crocodile serum were used to determine its quantitative inhibitory effect and mechanism of inhibition against the HIV-1 reverse transcriptase activity. A colorimetric enzyme immunoassay that incorporates digoxigenin and biotin-labeled dUTP onto DNA were used for the assay (Roche Version 13.0, 2010). In addition, the kinetic activity of HIV-1 Reverse transcriptase was studied to determine the mechanism of enzyme inhibition by Philippine crocodile serum. A minimum of 2 mL serum sample from each of the seven (7) purposively selected Crocodylus mendoremsis were collected at the Animal clinic of the Davao Crocodile Park facility. The results showed that the HIV-1 Reverse transcriptase activity was inhibited by high as 92.93±0.72% (CM-07) at 0.5 vol/vol % and as low as 4.66±20.76 at 4 vol/vol % (CM-04) of Philippine crocodile serum. The average half maximal inhibitory concentration (IC 50) of the seven crocodile serum samples was 1.47% vol/ vol.The 92.93±0.72% inhibition of 0.5% vol/vol of Philippine crocodile serum has almost the same inhibition as the positive control Nevirapine (92.64±0.20%). The inhibition kinetics of the Philippine crocodile suggests a potential novel HIV-1 Reverse Transcriptase inhibitor.

Anti-HIV-1 Activity of Crocodylus mindorensis (Philippine crocodile) serum in cell-free and cell-associated virus interactions to human peripheral blood mononuclear cells

“…This means that the inhibitory activity decreases as the Philippine crocodile serum concentration increases. This is similar to the previous study of Hinay and Sarol (2018), were the result could be explained by the presence of an “inhibitor of the inhibitor” in the crocodile serum. The best candidate inhibitor present in the crocodile serum with putative inhibition activity to HIV-1 is the Antimicrobial Peptides (AMPs) (Finger & Isberg, 2012).…”

“…The best candidate inhibitor present in the crocodile serum with putative inhibition activity to HIV-1 is the Antimicrobial Peptides (AMPs) (Finger & Isberg, 2012). This is also supported by the study of Hinay and Sarol (2018) where Philippine crocodile serum concentration showed in vitro inhibition of HIV-1 Reverse transcriptase by high as 92.93±0.72% at 0.5 vol/vol % (Hinay and Sarol, 2018). Moreover, another putative component of the Philippine crocodile serum as described by Okada and colleagues provided a good support that the complement fragment C5a, an anaphylotoxin and a small peptidase product during complement activation, has a 30% HIV-1 Reverse transcriptase inhibitory activity (Okada, et al, 2011).…”

“…The HIV-1 RT activity after 21 days was 0.5928 ng/well. On the other hand, a baseline Philippine crocodile serum concentration of 0.5% vol/vol was used based on the study conducted by Hinay and Sarol (2018) and the cell viability results showed no cell reduction of mononuclear cells after 72 hours incubation.…”

“…The HIV-1 reverse transcriptase activity after 21 days was 0.5928 pg/well. Moreover, a baseline Philippine crocodile serum concentration of 0.5% vol/vol was used based on the previous study conducted by Hinay and Sarol (2018) and the cell viability results showed no cell reduction of mononuclear cells after 72 hours incubation. The inhibitory activity of the Philippine crocodile serum at 0.5% and 0.25% vol/vol concentrations inhibited 65.68±2.93% and 69.92±0.45% respectively in post-infection interactions.…”

Anti-HIV-1 Activity of Crocodylus mindorensis Philippine Crocodile Serum in Cell-free and Cell-associated Virus Interactions to Human Peripheral Blood Mononuclear Cells

“…The HIV-1 RT activity after 21 days was 0.5928 ng/ well. On the other hand, a baseline Philippine crocodile serum concentration of 0.5% vol/vol was used based on the study conducted by Hinay and Sarol [9] and the cell viability results showed no cell reduction of mononuclear cells after 72 hours incubation.…”