Ethanol injection

A disadvantage of the ethanol injection method to produce SUV is the need to use a low lipid concentration, resulting in a low encapsulation efficiency of the aqueous phase. The dispersions can be concentrated by ultracentrifugation, ultrafiltration, or removal of water by evaporation. 

Foradada and Estelrich [3.63] studied the encapsulation of thioguanine (TG) in three types of liposomes produced by extrusion, ethanol injection and dehydration-rehydration vesicles. The entrapment has been examined at three different concentrations (1, 0.1 and 0.01 mM) and at three different pH values (4.7,7.4 and 9.2). The dehydration-rehydration vesicles were found to be the optimum method to encapsulate TG, independent of the pH value. At pH 4.7, 12 mmol/mol of lipid were entrapped, while with the other methods a maximum of 3 mmol/mol of lipid has been achieved. The authors related this behavior to the formation of hydrogen bridges between the TG and the liposomes. 

Rh was loaded on the support by impregnation. Among the catalysts tested in the SRE reaction, Rh/Ce02-Zr02 was the best producing 5.8 moles of H2/mole of ethanol injected and a high C02/CO ratio around 35 at 450 °C. 

The mixed liposomal solutions were prepared by the ethanol-injection method(13) in order to obtain completely transparent solutions. It is interesting to note that miscibility of the photochromic amphiphiles with DPPC depend on the position of bulky azobenzene. If azobenzene is incorporated close to the end of long alkyl chain, a stable mixed bilayer state cannot be formed. On the other hand, when the azobenzene moiety is located near the head group or at the center of the hydrocarbon tail, the azobenzene amphiphiles are successfully incorporated into the bilayer membrane. No individual micelle formation nor phase separation in the bilayer was observed at 25 °C by absorption spectroscopy. However, the microstructure of the mixed liposomes depends on the type of azobenzene amphiphiles. 

The procedure chosen for the preparation of lipid complexes of AmB was nanoprecipitation. This procedure has been developed in our laboratory for a number of years and can be applied to the formulation of a number of different colloidal systems liposomes, microemulsions, polymeric nanoparticles (nanospheres and nanocapsules), complexes, and pure drug particles (14-16). Briefly, the substances of interest are dissolved in a solvent A and this solution is poured into a nonsolvent B of the substance that is miscible with the solvent A. As the solvent diffuses, the dissolved material is stranded as small particles, typically 100 to 400 nm in diameter. The solvent is usually an alcohol, acetone, or tetrahydrofuran and the nonsolvent A is usually water or aqueous buffer, with or without a hydrophilic surfactant to improve colloid stability after formation. Solvent A can be removed by evaporation under vacuum, which can also be used to concentrate the suspension. The concentration of the substance of interest in the organic solvent and the proportions of the two solvents are the main parameters influencing the final size of the particles. For liposomes, this method is similar to the ethanol injection technique proposed by Batzii and Korn in 1973 (17), which is however limited to 40 mM of lipids in ethanol and 10% of ethanol in final aqueous suspension. 

In the procedure utilized by Oberholzer and Luisi (2002) all ingredients are added to a solution in which the vesicles are being formed by the ethanol injection method,... 

Gimatecan)-containing liposomes prepared by the ethanol injection method. J. Lipos. Res., 14, 87-109. 

When 30 % of ethanol (20 ml/kg) was administrated by the intravenous injection 10 min prior to tetanus blocked the induction of LTP. By the oral CSE (125 and 250 mg/kg) administrations 20 min prior to ethanol injection, the LTP-blocking effect of intravenously injected ethanol was significantly attenuated dose-dependently Fig. (8-A). Fig. (8-B) indicated the dose-dependency of the influence of CSE on the LTP-blocking effect of ethanol. The effect of CSE was also observed when ethanol was injected intracerebroventricularly [13],... 

Perhaps the simplest solvent dispersion method is that developed by Batzri and Korn (1973). Phospholipids and other lipids to be a part of the liposomal membrane are first dissolved in ethanol. This ethanolic solution is then rapidly injected into an aqueous solution of 0.16 M KC1 using a Hamilton syringe, resulting in a maximum concentration of no more than 7.5% ethanol. Using this method, single bilayer liposomes of about 25-nm diameter can be created that are indistinguishable from those formed by mechanical sonication techniques. The main disadvantages of ethanolic injection are the limited solubility of some lipids in the solvent (about 40 mM for phosphatidyl choline) and the dilute nature of the resultant liposome suspension. However, for the preparation of small quantities of SUVs, this method may be one of the best available. 

Stano, P, Bufali, S., Pisano, C., Bucci, F., Barbarino, M., Santaniello, M., Carminati, P, and Luisi-Pier, L. (2004). Novel camptothecin analogue (gimatecan)-containing liposomes prepared by the ethanol injection methodJ. Liposome Res., 14, 87-109. 

Erythrocytes from ethanol-injected fish permeated more water than those from normal fish. Water permeation was significantly enhanced with Merc cor 30 and Nux vom 30 as compared to the control. RBCs from fish pretreated with Nux vom 30 inbibed more water in in vitro treatments than those from fish pre-treated with Ethanol 30. Since aquaporins are mainly responsible for water transport through the plasma membrane of red blood cells it is thought that potentized drugs such as Merc cor 30 and Nux vom 30 acted upon these proteins and facilitated water influx into the cells (Sukul et al., 2003). 

The separation of cimetidine and its metabolites is usually carried out by extraction of the biological medium with 1-octanol fran an aqueous alkaline pH solution followed by mixing, addition of an internal standard and centrifugation. The extraction with octanol is repeated and the combined extracts are re-extracted with dilute hydrochloric acid. The aqueous acid solution is then separated, ethanol is added and mixed. This is then followed by saturating the mixture with a large amount of potassium or sodium carbonate to "salt out" the ethanol layer which contains the cimetidine and its metabolite, the sulfoxide. Several different internal standards have been used Metiamide, 1-methyl-3-[2-[[(5-methyl-imidazole-4-yl) -methyl] thio]ethyl]-2-thiourea,19 31 39 (N-cyano-N1-methy1-N"-(3-(4-imidazolyl)-propyl)guanidine32, and 13-hydroxy-theophylline. 0 After extraction the samples are either evaporated to dryness and reconstituted with a known amount of ethanol, injected directly or dissolved in the mobile phase for the HPLC analysis. 

Ethanol Injection Small unilamellar vesicles (with diameter of 30 nm) can be prepared with the ethanol injection technique [128], Lipids are dissolved in ethanol and injected rapidly in the aqueous solution under stirring (final concentrations up to 7.5% (v/v) ethanol can be applied).The method is very easy, having the advantage of avoiding chemical or physical treatment of lipids. However, there is an extra step to remove ethanol and the concentration of vesicles produced is rather low. Also encapsulation of hydrophilic drugs is also low, due to the high volumes used. 

Ether Injection The general principle of this method is the same as ethanol injection. The only difference is that the lipid is injected slowly in the aqueous solution that is warm [129], Furthermore, the concentrations used in this case are somewhat higher (up to 10mM) compared to the ethanol injection approach. 

The manufacturing of nanosized liposomes can be performed using the methods mentioned above. However, the small size of nanoliposomes is difficult to achieved by methods such as film hydration. Molecular self-assembly occurs in the injection method, and then the size and morphology of obtained liposomes can be well controlled. In fact, liposomes that result from the injection method are uniform and small enough, to the nanoscale, and usually SUVs are obtained. Because of the very low toxicity of ethanol, the ethanol injection method is usually used and is described as follows to show the process of manufacturing liposomes [50]. A scale-up manufacturing process of the ethanol injection method has been established [80-82], The obtained liposome size is mostly less than 300 nm ... 

Wagner, A., Vorauer-Uhl, K., Kreismayr, G., and Katinger, H. (2002), The crossflow injection technique—An improvement of the ethanol injection method, J. Liposome... 

The Step 2 product was packed into an HPLC column and IM solution pentaethylene hexamine and 0. IM acetic acid in anhydrous ethanol injected into the column. After 2 hours a 0.6M sodium borohydride solution in anhydrous ethanol was injected into the column. After an additional hour unreacted reagents were flushed from the column. The resulting polypentaethylene hexamine silica was able to hold about 800 pmol copper per gram of silica gel. 

Giorgio, A., Tarantino, L., de Stefano, G., Francica, G., Marinlello, N., Farella, N., Perrotta, A., Aloisio, V., Esposito, F. Hydatid liver cyst. An 11-year experience of treatment with percutaneous aspiration and ethanol injection. J. Ultrasound Med. 2001 20 729 - 738... 

Isobe, H., Fukai, T., Iwamoto, H., Satoh, M., Tokumatsu, M., Sakai, H., Andoh, B., Sakamoto, S., Nawata, H. Liver abscess complicating intratumoral ethanol injection therapy for HCC. Amer. J. Gastroenterol. 1990 85 1646-1648... 

Okada, S., Aoki, K., Okazaki, N., Nose, H., Yoshimori, M., Shimada, K., Yamamoto, J., Takayama, T., Kosuge, T., Yamasaki, S., Takayasu, K., Moriyama, N. Liver abscess after percutaneous ethanol injection (PEI) therapy for hepatocellular carcinoma. A case report. Hepato-... 

Solinas, A., Erbella, G.S., Distrutti, E., Malaspina, C., Fiorucci, St., Clerici, C., Bassotti, G., Morelli, A. Abscess formation in hepatocellular carcinoma complications of percutaneous ultrasound-guided ethanol injection. J. Clin. Ultrasound 1993 21 531-533... 

Fig. 37.9 Subcapsular HCC in liver cirrhosis mostly necrotic following ethanol injection...

Bartolozzi, C., Lencioni, R., Ricci, P., Paolicchl, A., Rossi, P., Passa-riello, R. Hepatocellular carcinoma treatment with percutaneous ethanol injection evaluation with contrast-enhanced color Doppler US. Radiology 1998 209 387- 393... 

Kawamoto, C., Ido, K., Isoda, N., Nagamine, N., Hozumi, M., Ono, K., Nakazawa, Y., Sato, Y., Kimura, K. Prognosis of small hepatocellular carcinoma after laparoscopic ethanol injection. Gastrointest. Endosc. 1999 50 214- 220... 

Khan, K.N., Yatsuhashi, H., Yamasaki, K., Yamasaki, M., Inoue, O., Koga, M., Yano, M. Prospective analysis of risk factors for early intra-hepatic recurrence of hepatocellular carcinoma following ethanol injection. J. Hepatol. 2000 32 269 - 278... 

Koda, M., Murawaki, Y., Mitsuda, A., Ohyama, K., Horie, Y., Suou, T., Kawasaki, H., Ikawa, S. Predictive factors for intrahepatic recurrence after percutaneous ethanol injection therapy for small hepatocellular carcinoma. Cancer 2000 88 529-537... 

Hepatocellular carcinoma and cirrhosis in 746 patients long-term residts of percutaneous ethanol injection. Radiology 1995 197 101-108... 

Livraghi, T., Goldberg, S.N., Lazzaroni, F., Meloni, F., Solbiati, L., Gazelle, G.S. Small hepatocellular carcinoma Treatment with radiofrequency ablation versus ethanol injection. Radiology 1999 210 655-661... 

Mazzanti, R., Arena, U., Pantaleo, P., Antonuzzo, L., Cipriani, G., Neri, B., Giordano, C., Lanini, F., Marchetti, S., Gentilini, P. Survival and prognostic factors in patients with hepatocellular carcinoma treated by percutaneous ethanol injection A 10-year experience. Can. J. Gastroenterol. 2004 18 611—618... 

Miki, K., Makuuchi, M., Taka.yama, T., Matsukura, A., Minagawa, M., Kubota, K., Hirata, M. Peritoneal seeding of hepatocellular carcinoma after ethanol injection therapy. Hepato-Gastroenterol. 2000 47 1428-1430... 

Pompili, M., Rapacdni, G.L., de Luca, F., Caturelll, E., Astone, A., Siena, D.A., Villanl, M.R., Grattagliano, A., Cedrone, A., Gasbarrini, G. Risk factors for intrahepatic recurrence of hepatocellular carcinoma in cirrhotic patients treated by percutaneous ethanol injection. Cancer 1997 79 1501-1508... 

Yamamoto, K., Masnzawa, M., Karo, M., Kurosawa, K., Kaneko, A., Ishlda, H., Imamnra, E., Park, N.J., Shirai, Y., Fnjimoto, K., Michida, T., Hayashl, N., Breda, M. Evaluation of combined therapy with chemoemholization and ethanol injection for advanced hepatocellular carcinoma. Semin. Oncol. 1997 24 (Suppl. 6) 50—55... 

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