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  • Yoda1 br Evaluation of the release characteristics of CPT

    2022-05-22


    Evaluation of the release characteristics of CPT from the DSPE-NLBs bore a strong resemblance to that obtained from the CHO-NLBs in both a physiologic and tumoral pH release medium. This observation was attributed to lower stability of the DSPE-NLBs, due to the less anionic surface charge, as well as the higher permeability of DSPE-NLBs through the lipid membrane. Release of CPT from DSPE-NLBs was faster over the first 8 h of evaluation. The cumulative release of CPT from the DSPE-NLBs at physiological and tumoral pH was approximated to be 50% and 58%, respectively.
    SB is a naturally occurring polyphenol antioxidant extracted from milk thistle (Silybum marianum) and has an excellent safety profile, being well tolerated at considerably high doses following different routes of administration [18–21]. It is also known for its anti-cancer properties where the mechanism of its anti-tumor effect has been at-tributed to factors such as the promotion of cellular antioxidant defense mechanisms, angiogenesis antagonism by a reduction in VEGF secre-tion, induction of apoptosis and antiproliferative effects due to Yoda1 arrest [18,20,22]. SB, however, has a limited aqueous solubility and undergoes metabolism by phase II conjugation. The incorporation of silibinin in the formulated NLB-DDS was therefore undertaken to enhance the cytotoxic activity of the formulations and provide a means of effective delivery of this poorly soluble phytochemical. Incorporation of silibinin into the formulated NLB-DDS revealed a size of 137.56 nm and 93.65 nm, a zeta potential of -27.58 mV and -8.24 mV and a drug incorporation efficiency of 65.59% and 52.75% for the CHO-NLB and DSPE-NLB systems respectively.
    The release pattern of CPT from CHO-NLBs containing SB (CHO-NLB + SB) demonstrated no outstanding differences to that of SB for-mulations for the first 10 h, except for an evident burst release of CPT over the first hour (Fig. 2A). A similar burst release of SB was observed over this period, displayed in Fig. 2B, suggesting association of both compounds to a certain degree with the surface of the NLBs. Further-more, the presence of the additional SB compound may have altered the surface tension of the formulated CHO-NLBs, leading to the initial burst release of both CPT and SB. From 10 h the release of CPT from CHO-NLB + SB is approximately 7–9% higher than that observed for CHO-NLB without SB. An effect of the different pH of release medium (7.4 and 6.0) employed during analysis only became evident after 10 h, when the release of CPT at pH 6.0 appeared to be slightly higher than that at pH 7.4. However, the effect of tumoral pH on the release characteristics of CPT from CHO-NLB + SB was still considered negli-gible. A cumulative CPT release of 82–86% was achieved for CPT from CHO-NLB + SB over the 24-hour investigation.
    3.5. Cellular uptake of nano-lipobubbles
    Formulated CHO- and DSPE-NLBs demonstrated a time-dependent cellular uptake, which was more pronounced in the CHO-NLBs. The distinct increase in fluorescence intensity evident in the fluorescence micrographs in the upper panel of Fig. 3 highlights the increase in CHO-NLB uptake by the A2780 cells 30 min post-treatment relative to that depicted 10 min post-treatment. Cellular uptake of the DSPE-NLBs was markedly higher and faster than observed for CHO-NLBs, which was attributed to the smaller size of DSPE-NLBs. Localization of both CHO-
    Fig. 3. Qualitative evaluation of cellular uptake of SRB-labeled CHO- and DSPE-NLBs at 10 and 30 min post-treatment.
    and DSPE-NLBs highlight the potential for released CPT to be trans-ported to the cell nucleus, which is the target for antineoplastic activity.
    3.5.1. Assessment of cytotoxicity by flow cytometry