• Journal of Pharmaceutical Investigation
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• 제34권1호
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• pp.1-14
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• 2004

A promising approach to the development of a new single-step vaccine, which would eliminate the requirement for multiple injections, involves the encapsulation of antigens into microspheres. Biodegradable poly(lactide-co-glycolide) (PLGA) microspheres gave us a bright insight for controling antigen release in a pulsatile fashion, thereby mimicking two or tree boosting injections. However, in spite of the above merits, the level of immunization induced by a single-shot vaccination is often lower tan two doses of alum-adsorbed antigen. Therefore, optima modification of the microsphere is essential for the development of single-step vaccines. In the review, we discuss the stability of antigen in microsphere, safety and non-toxic in human and encapsulation technology. Also, we attempted to outline relevant physicochemical properties on the immunogenicity of microsphere vaccine and attainment of pulsatile release pater by combination of different microsphere, as well as to analyze immunological data associated with antigen delivery by microsphere. Although a lot of variables are related to the optimized microsphere formulation, we could conclude that judicious choice of proper polymer type, adjustment of particles size, and appropriate immunization protocol along with a suitable adjuvant might be a crucial factor for the generation of long-lasting immune response from a single-step vaccine formulation employing PLGA microsphere.

• 약학회지
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• 제44권6호
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• pp.511-520
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• 2000

Various bupivacaine-loaded microspheres were prepared from poly (d,l-lactide) (PLA) or poly (d,l-lactic-co-glycolide) (PLGA) by a solvent evaporation method for the sustained release of drug. PLA and PLGA microspheres were prepared by w/o/w and w/o/o multiple emulsion solvent evaporation, respectively. The effects of process conditions such as emulsification speed, emulsifier type, emulsifier concentration and internal/external phase ratio on the characteristics of microspheres were investigated. The prepared microspheres were characterized for their drug loading, size distribution, surface morphology and release kinetics. Drug loading efficiency was higher in the microspheres prepared by w/o/o multiple emulsion than that by w/o/w multiple emulsion method, because the solubility of bupivacaine HCI was decreased in oil phase compared with water phase. The prepared microspheres had an average diameter between 1 and $2\;{\mu}M$ in all conditions of two methods. In morphology studies the PLA microspheres showed an irregular shape and smooth surface, but PLGA microspheres had a spherical shape and smooth surface. The release pattern of the drug from microspheres was evaluated on the basis of the burst effect and the extent of the release after 24h. The in vitro release of bupivacaine HCl from microspheres showed a large initial burst release and $60{\sim}80%$ release within one day in all conditions of two methods. The extents of the burst release against PLA and PLGA microspheres were $30{\sim}50%$ and $50{\sim}80%$ within 20min, respectively. This burst release seems to be due to the smaller size of microspheres and the solubility of drug in water.

• Journal of Pharmaceutical Investigation
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• 제31권3호
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• pp.191-196
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• 2001

Biodegradable polymeric microspheres were studied for their usefulness as carriers for the delivery of vaccine antigens. However, protein antigen could be denatured during microencapsulation processes due to the exposure to the organic phase and stress condition of cavitation and shear force. Therefore this study was carried out to re-evaluate the degree of protein denaturation during microencapsulation with poly(lactide-co-glycolide) (PLGA) copolymer. PLGA microspheres containing ovalbumin (OVA), prepared by W/O/W multiple emulsification method, were suspended in pH 7.4 PBS and incubated with shaking at $37.5^{\circ}C$. Drug released medium was collected periodically and analyzed for protein contents by micro-BCA protein assay. In order to evaluate the protein integrity, release medium was subjected to the analyses of SDS-PAGE and size exclusion chromatography (SEC). And enzyme-linked immunosorbent assay (ELISA) was introduced to measure the immunoreactivity of entrapped OVA and to get an insight into the three-dimensional structure of epitope. The structures of entrapped protein were not affected significantly by the results of SDS-PAGE and SEC. However, immunoreactivity of released antigen was varied, revealing the possibility of protein denaturation in some microspheres when it was evaluate by ELISA method. Therefore, in order to express the degree of protein denaturation, antigenicity ratio (AR) was obtained as follows: amount of immunoreactivity of OVA/total amount of OVA released ${\times}100(%)$. ELISA method was an efficient tool to detect a protein denaturation during microencapsulation and the comparison of AR values resulted in more accurate evaluation for immunoreactivity of entrapped protein.

• 대한수의학회지
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• 제44권1호
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• pp.113-120
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• 2004

1% iodophor loaded microspheres of PLGA (Poly[DL-Lactide-co-Glycolide]) were prepared by solvent evaporation method and were applied to the cows on dry period for evaluating it's preventive effects on intramammary infections. The morphology of the microspheres were evaluated using scanning electron microscopy and their releasing patterns were investigated. On investigating idophor releasing patterns of the microsphere, burst releasing pattern was detected until 2 days after in vitro incubation and sustained releasing was observed until 4 weeks. In field trial of teat dipping solution containing idophor loaded microspheres in dry cows showed significant preventive effects of intramammary infection caused by S. aureus, S. agalactiae, coagulase negative Staphylococci and coliform bacteria (p<0.05).

• Journal of Pharmaceutical Investigation
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• 제36권4호
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• pp.245-251
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• 2006

The novel microsphere blending and multiple emulsion method by single process was tried to prepare sustained release microspheres which release a physiologically active substance for long periods of time. A drug was separately dissolved in each of two or more oils containing biodegradable polymers to give the primary oil phases. The primary oil phases were dispersed in single aqueous phase in succession. From the drug-dispersed solution, the organic solvent was removed to produce microspheres. The accelerated drug release from the microsphere formulation prepared by single process through the multiple emulsion method was very similar to a physical blending of separately prepared microspheres using the same polymers. But long term release was not same. In this study, leuprorelin acetate loaded poly(lactide-co-glycolide) microsphere formulation for one-month delivery was developed by the multi-emulsion method followed by solvent extraction/evaporation method.

• 대한약학회:학술대회논문집
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• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2
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• pp.417.2-418
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• 2002

The rhGH-loaded PLGA microsphere formulation was prepared using a double emulsion process from hydrophilic 0:50 poly(D.L-lactide-co-glycolide) (PLGA) polymers. To investigate the sustained efficacy of this formulation, ts pharmacodynamic characteristics were analyzed. It showed particle size of ca 53.1 $\mu\textrm{m}$ with high drug ncorporation efficiency and it was sucutaneously administrated to hypophysectomized rats and whole body rowth responses of this formulation were compared to those of the different dosing patterns of rhGH. (omitted)

• 대한약학회:학술대회논문집
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• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2
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• pp.424.2-424.2
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• 2002

The in vivo release characteristics of rhGH-loaded PLGA microsphere prepared using a double emulsion process from hydrophilic 50:50 poly(D.L-lactide-co-glycolide) (PLGA) polymers were analyzed. This formulation showed particle size of ca 53.1$\mu\textrm{m}$ with high drug incorporation efficiency. To investigate in vivo release kinetics without the interference of formation of antibodies to rhGH in the experimental animals, the animals were immunosuppressed by treatment with Cyclosporin. (omitted)

• Journal of Pharmaceutical Investigation
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• 제34권5호
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• pp.369-377
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• 2004

The objective of this study was to investigate the patterns of protein leaching to an external phase during an ethyl acetate-based, double emulsion microencapsulation process. An aqueous protein solution (lactoglobulin, lysozyme, or ribonuclease; $W_1$) was emulsified in ethyl acetate containing poly-d,l-lactide-co-glycolide 75:25. The $W_1/O$ emulsion was transferred to a 0.5% polyvinyl alcohol solution saturated with ethyl acetate $(W_2)$. After the double emulsion was stirred for 5, 15, 30, or 45 min, additional 0.5% polyvinyl alcohol $(W_3)$ was quickly added into the emulsion. This so-called quenching step helped convert emulsion microdroplets into microspheres. After 2-hr stirring, microspheres were collected and dried. The degree of protein leaching to $W_2$ and/or $W_3$ phase was monitored during the microencapsulation process. In a separate, comparative experiment, the profile of protein leaching to an external phase was investigated during the conventional methylene chloride-based microencapsulation process. When ethyl acetate was used as a dispersed solvent, proteins continued diffusing to the $W_2$ phase, as stirring went on. Therefore, the timing of ethyl acetate quenching played an important role in determining the degree of protein microencapsulation efficiency. For example, when quenching was peformed after 5-min stirring of the primary $W_1/O$ emulsion, the encapsulation efficiencies of lactoglobulin and ribonuclease were $55.1{\pm}4.2\;and\;45.3{\pm}7.6%$, respectively. In contrast, when quenching was carried out in 45 min, their respective encapsulation efficiencies were $39.6{\pm}3.2\;and\;29.9{\pm}11.2%$. By sharp contrast, different results were attained with the methylene-chloride based process: up to 2 hr-stirring of the primary and double emulsions, less than 5% of a protein appeared in $W_2$. Afterwards, it started to partition from $W_1\;to\;W_2/W_3$, and such a tendency was affected by the amount of PLGA75:25 used to make microspheres. Different solvent properties (e.g., water miscibility) and their effect on microsphere hardening were to be held answerable for such marked differences observed with the two microencapsulation processes.

• 폴리머
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• 제29권6호
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• pp.543-548
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• 2005

이중층 미립구는 단일층 미립구에 비해서 낮은 초기 방출량과 국소 지항성, 및 약물 방출량 제어 등의 장점 을 갖고 있다. 그러나 포접 방식이 까다롭고 2단계 이상의 제조 과정이 필요하며 특히 미립구 제조 시 크기 조절이 어렵다는 단점을 갖고 있다. 따라서 본 연구에서는 락타이드글리콜라이드 공중합체(PLGA)와 덱스트란의 서로 다른 고분자를 초고주파 분쇄 유무에 따른 수중유형(O/W) 용매 증발법을 이용하여 이중층 미립구를 제조하였다. 또한 PLGA의 농도에 따라 미립구 크기의 변화를 연구하였다. 이중층 미립구는 전자주사현미경, 동초점 형광 레이저현미경(CFLM), 캠스코프를 이용하여 조사하였다. 제조된 이중층 미립구의 외부층이 매끄러운 구형의 형태를 나타냄을 확인할 수 있었으며, 절단시 내부층과 외부층의 형태를 확실히 구분할 수 있었다. 이에 외부층과 내부층의 구성 물질을 확인하고자 플루오르신-5-이소시아네이트-덱스트란(FITC-덱스트란)을 이용해 CFLM을 관찰한 결과 형광을 띠는 덱스트란으로 구성된 내부층과 형광을 띠지 않는 PLGA 외부층을 관찰하였다. 또한 PLGA의 함량에 따른 미립구의 크기는 전체적으로 증가하는 경향을 확인하였다. 이와 같은 결과로부터 비교적 간단한 수중유형 용매 증발법을 이용하여 PLGA와 덱스트란의 이중층 미립구의 제조가 가능함을 확인할 수 있었다.

• 폴리머
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• 제33권1호
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• pp.26-32
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• 2009

골 결손 치료를 위해 생분해성 고분자인 PLGA에 골다공증 치료제인 이프리플라본(IP)을 함유한 미립구를 O/W 유화 용매 증발법으로 제조하였으며, 생체외 방출실험에서 IP 방출량은 HPLC로 분석하였다. SEM을 이용하여 미립구에 부착된 세포의 거동을 확인하였으며, IP가 세포에 미치는 독성평가는 CCK-8 분석방법을 이용하여 측정하였다. 골 형성 지표인 ALP 활성도를 측정하였으며, 배양된 BMSCS의 골세포로의 표현형을 확인하기 위하여 RT-PCR을 수행하였다. 방출결과에서 IP 방출은 거의 40일 이상으로 지속적이었으며, IP를 함유한 미립구에서의 세포의 부착, 성장 등이 잘 이루어짐을 확인하였고, ALP 활성도 및 RT-PCR 분석결과에서도 단독의 PLGA 미립구보다 IP를 함유한 미립구에서의 값이 더 높은 것을 확인할 수 있었다. 본 실험결과를 바탕으로 향후 서방성 제제화에 있어서 IP/PLGA 미립구를 응용하게 되면 국소지향성 골분화 주사용 지지체로서 중요한 역할을 할 것으로 보인다.