Study of gas bubbles stabilized by surfactants for use as ultrasound contrast agents and drug carriers
Authors:   , Methachan Boriphat , Wheatley Margaret A. 
Published: July 2021

It has been well established that the vasculature in a tumor is more 'leaky' than that in normal tissue. The cutoff pore size is in the range of 380 and 780 nm depending on the tumor model and stage. It would be useful to fabricate an ultrasound contrast agent (UCA) that is smaller than the cut off pore size of a tumor allowing it to flow into the tumor. This would facilitate better perfusion yielding better tumor imaging and lead to a new and more effective way to deliver the chemotherapeutic drug for cancer treatment. One of UCAs fabricated our lab was gas bubbles stabilized by surfactants. Specifically, perfluorocarbon (PFC) gas bubbles have been stabilized by the mixture of two surfactants. Two surfactant stabilized agents were developed: 1) a PFC gas bubble contrast agent, ST68, stabilized with Span60 and Tween 80 with a mean diameter of 1.88±0.16 [mu]m and 2) SE61, with a Span60, d-[alpha] tocopheryl polyethylene glycol 1000 succinate, or TPGS shell and comprised of gas bubbles with a mean diameter of 1.31±0.20 [mu]­m. These agents provided 20-30 dB ultrasound enhancements and the stability was dependent on the particle size that stabilized the bubbles. A new model was proposed to explain a previously unexplained observation, i.e., the fact that only combinations of solid Span with Tween (a liquid) can form microbubbles, liquid Spans do not. Various studies were conducted which indicate that the bubbles stabilized by solid particles is a realistic possibility. These included the behaviors of the bubbles as they dried, the fact that larger Span particles produced more robust bubbles and preliminary scanning electron microscopic evidence. A substantial number of nanobubbles were fabricated for SE61 compared with ST68. These nanobubbles still retained good echogenicity, but they were less stable (1.75±0.24 min) as compared to the microbubble (16.45±1.56 min). ST68 with a bubble diameter ranging from 955 nm to 2.67 [mu]m and SE61 with bubble sizes ranging from 342 nm to 900 nm were used to see if the smaller sized bubbles would yield better ultrasound imaging for tumor perfusion deeper into the tumor bed. Power-Doppler ultrasound imaging was used in vivo to analyze the tumor perfusion for both agents in an animal (rat) study. Tumor coverage and time were plotted and analyzed by comparing parameters in wash-in and wash-out phases. The results showed that both agents gave good tumor coverage. However, SE61 provided better perfusion imaging than ST68 and took longer to clear out of the tumor. It should be noted that Delta-projection is better in portraying the vasculature of the tumor as compared to Power-Doppler. By using Nile Red, a hydrophobic fluorescent solvatochromic probe, we showed that a hydrophobic drug could intercalate into the hydrophobic portion of the surfactants in the bubble shell to make drug delivery possible. The subsequent experiments showed that Paclitaxel, a promising anti-tumor agent with poor water solubility, intercalated into the bubbles at the concentration around 3.8 [mu]g/ml. This concentration was close to 100 times higher than the suggested effective concentration from other studies.