Preparation and in vitro/in vivo characterization of enteric-coated nanoparticles loaded with the antihypertensive peptide VLPVPR

Abstract

Our previous study revealed that the peptide Val-Leu-Pro-Val-Pro-Arg (VLPVPR), which was prepared using deoxyribonucleic acid recombinant technology, effectively decreased the blood pressure of spontaneous hypertensive rats; however, the effect only lasts 6 hours, likely due to its low absorption in the gastrointestinal tract. To overcome this problem, the purpose of this study was to characterize (methoxy-polyethylene glycol)-b-poly(D,L-lactide-co-glycolide)-b-poly(L-lysine) nanoparticles as in vitro and in vivo carriers for the effective delivery of VLPVPR. In our study, the VLPVPR nanoparticles were prepared using a double emulsion method, coated with Eudragit S100, and freeze-dried to produce enteric-coated nanoparticles. The optimized parameters from the double emulsion method was obtained from orthogonal experiments, including drug loading (DL) and encapsulated ratio (ER) at 6.12% and 86.94%, respectively, and the average particle size was below 100 nm. The release experiment demonstrated that the nanoparticles were sensitive to pH: almost completely released at pH 7.4 after 8 hours, but demonstrated much less release at pH 4.5 or pH 1.0 in the same amount of time. Therefore, the nanoparticles are suitable for enteric release. In vivo compared with the untreated group, the medium and high doses of orally administered VLPVPR nanoparticles reduced blood pressure for more than 30 hours, demonstrating that these nanoparticles have long-lasting and significant antihypertensive effects in spontaneously hypertensive rats.

Keywords: Val-Leu-Pro-Val-Pro-Arg peptide; antihypertensive peptide; enteric-coated; in vivo studies; mPEG-PLGA-PLL; nanoparticle.

Figure 1

Typical SEM images of mPEG-PLGA-PLL nanoparticles coated with Eudragit S100; bar, 200 nm. Abbreviations: mPEG-PLGA-PLL, (Methoxy-polyethylene glycol)-b-poly(D,L-lactide-co-glycolide)-b-poly(L-lysine); SEM, scanning electron microscopy.

Figure 2

Particle size of nanoparticles prepared using the double emulsion method.

Figure 3

Cumulative release of free VLPVPR in PBS (pH =7.4). Abbreviations: h, hours; PBS, phosphate buffered saline; VLPVPR, Val-Leu-Pro-Val-Pro-Arg.

Figure 4

Cumulative release of three enteric-coated nanoparticles in different pH media. Notes: (A) Cumulative release of enteric-coated nanoparticles (mPEG-PLGA-PLL:Eudragit S100 100:3.5) in pH 1.0, 4.5, and 7.4 media. (B) Cumulative release of enteric-coated nanoparticles (mPEG-PLGA-PLL:Eudragit S100 100:7) in pH 1.0, 4.5, and 7.4 media. (C) Cumulative release of enteric-coated nanoparticles (mPEG-PLGA-PLL:Eudragit S100 100:10.5) in pH 1.0, 4.5, and 7.4 media. Abbreviations: h, hours; mPEG-PLGA-PLL, (Methoxy-polyethylene glycol)-b-poly(D,L-lactide-co-glycolide)-b-poly(L-lysine).

Figure 5

Effects of a single oral dose of mPEG-PLGA-PLL nanoparticles on SBP in SHR. Notes: n=8, $\overline{X}\pm \text{s}$. Letters indicate the level of significant difference from control: ^aP<0.01; ^bP<0.05. Abbreviations: AHP, antihypertensive peptides; mPEG-PLGA-PLL, (Methoxy-polyethylene glycol)-b-poly(D,L-lactide-co-glycolide)-b-poly(L-lysine); s, seconds; SBP, systolic blood pressure; SHR, spontaneously hypertensive rats.