In order to design an optimum liposome immunoadjuvant for tumor vaccines, we investigated the relationship between liposomal surface charge and adjuvant action. Positively charged multilamellar vesicles (MLVs) were taken up efficiently by macrophages, while negatively charged and neutral MLVs were hardly picked up. Consistent with this, positively charged MLVs containing soluble ovalbumin (OVA) functioned as a more potent inducer of antigen-specific cytotoxic T lymphocyte (CTL) responses and antibody production than negatively charged and neutral MLVs containing the same concentrations of antigens. Furthermore, the in vivo anti-tumor effects of variously charged liposomal antigens were examined using a Meth A tumor model and a crude butanol extract derived from Meth A (Meth A-CBE) as the tumor-associated antigen. Mice vaccinated with positively charged MLVs containing Meth A-CBE showed significant inhibition of Meth A tumor growth compared to mice vaccinated with Meth A-CBE alone or mice vaccinated with neutral or negatively charged liposomal Meth A-CBE. The injection of carrageenan into mice led to a significant loss of anti-tumor vaccinal effect of positively charged liposomal Meth A-CBE, which may be due to the inhibition of uptake and antigen presentation of liposomal antigens by macrophages as a result of a lack of macrophages in the immune site. Our results indicate that the positive charge on the surface of liposomes represents an important factor for enhancing their immunoadjuvancy in the induction of antigen-specific immune responses and vaccinal effects against tumors.

The effects of emulsifiers on the plasma disappearance of lipid emulsion (LE) were studied in rats. Lipophilic model drugs, oil violet and sudan III, were incorporated into LE which consisted of soybean oil and emulsifiers such as egg yolk lecithin (PC), a mixture of PC and sphingomyelin (SM) (PC+SM), or Pluronic F-127 (PLURONIC). The plasma disappearance of LE containing PLURONIC (PLURONIC-LE), as determined from the triglyceride concentration, was slower than that when using PC-LE or (PC+SM)-LE. The area under the plasma concentration curve (AUC) of PLURONIC-LE was about 4-6 times larger than that of the other LEs. The plasma disappearance of drugs was also slowest when the drugs were incorporated in PLURONIC-LE. However, the effect of emulsifier on the AUC of the drugs was smaller than that of the LEs, suggesting that drugs were rapidly released from LE in the plasma. The production rates of non-esterified fatty acid (NEFA) from LE by lipoprotein lipase were in the following order: PC-LE>(PC+SM)-LE>PLURONIC-LE, suggesting that plasma disappearance of LE became slower with the decreasing order in the hydrolysis rate of triglyceride. The release of drugs from LE was not significantly changed by the hydrolysis of triglyceride. The hepatic concentration of oil violet after intravenous administration of LE was time-dependently increased in the case of PC-LE and PLURONIC-LE, and the concentration for PC-LE was higher than that for PLURONIC-LE. Consequently, it was suggested that the plasma concentration and hepatic distribution of lipophilic drugs in LE can be controlled by emulsifiers.

The stability and degradation mechanism of disodium 2-O-octadecyl-5,6-di-O-sulfoascorbate (code name, CV-11464), which is a new drug for anti-myocardial infarction, were studied in an aqueous solution. CV-11464, an ascorbate derivative, was unstable in an aqueous solution at room temperature (26°), and was the most unstable in a buffer solution, pH 4.9. The drug in an aqueous solution apparently degraded in a pseudo first-order manner, in which two degradation processes, fast and slow rates, were found. These processes were determined to be oxidation and hydrolysis reaction, respectively. The two degradation products in the solution were detected at the retention times of 2.3 and 7.6 min by HPLC determination. The degradation product at the retention time of 7.6 min was stable and could be collected by separation using HPLC, whereas the degradation product of 2.3 min was so unstable that could not be collected separately by HPLC. The chemical structure of the product collected at 7.6 min was estimated by nuclear magnetic resonance spectroscopy as follows: oxidation of 3-OH, hydrolysis of the 5 and 6 sulfate ester bond and lactone-ring opening by hydrolysis.

The stabilization of disodium 2-O-octadecyl-5,6-di-O-sulfoascorbate (code name, CV-11464), which is a new drug for anti-myocardial infarction, was studied in an aqueous solution. CV-11464, an ascorbate derivative, was unstable in an aqueous solution and was decomposed by two degradation processes. Stabilization of the drug in the solution was attained by adding ascorbic acid and maltosyl-β-cyclodextrin (G₂-β-CyD). A possible mechanism of the stabilization is that the ascorbic acid reacted to antioxidize 3-OH of the drug and G₂-β-CyD interacted to interrupt hydrolysis of the 5,6-diesters and lactone ring.

The influence of the physicochemical properties of fatty suppository bases on the vaginal absorption of prasterone sulfate (PS) was investigated. The suppository bases tested were Witepsol^® H-15, H-35 and W-35, and the vaginal absorption of PS from these suppositories was studied using rats. The area under the plasma concentration-time curve (AUC) of PS obtained with H-15 suppository was higher than that observed using H-35 or W-35 suppository. No differences in the viscosity of suppository, PS permeability through vaginal membrane or the release of PS were detected among H-15, H-35 and W-35 suppositories. The spreading of base and drug of the H-15 suppository was superior to that of the H-35 or W-35 suppository. These results indicate that Witepsol^® H-15 is an appropriate base for the preparation of PS vaginal suppositories.