Fabrication, optimization, and digestion of a sulforaphane delivery system using an oil-in-water emulsion and complex coacervate = 수중유 에멀젼 및 복합 코아세르베이션을 이용한 설포라판 전달 시스템의 제조, 최적화 및 소화성 평가

다국어 초록 (Multilingual Abstract)

Sulforaphane (SFN), known for its antioxidant effects, is structurally unstable and easily degrades under the acidic conditions of the gastrointestinal tract. Promising delivery strategies must be developed to stabilize and protect lipophilic compounds such as SFN in acidic gastric environments. The objective of this study was to develop an SFN-loaded O/W emulsion coated through complex coacervation (SFN-ECC) to enhance stability and targeted intestinal delivery. The O/W emulsion was prepared by dissolving Tween 80 and ι-carrageenan in water for the aqueous phase and dissolving SFN in sunflower oil for the oil phase. Mung bean protein isolate (MPI) was subsequently mixed to form SFN-ECC. Response surface methodology was used in a two-stage process to optimize SFN-ECC. The initial stage optimized the key emulsion characteristics (stability, droplet size, and distribution), and the subsequent stage focused on encapsulation efficiency, particle size, and coacervate yield. This optimization yielded the following optimal parameters: 54% sunflower oil, 0.60% ι-carrageenan, 8,793 rpm homogenization speed, pH 3.71, and 1.17% MPI. The optimized SFN-ECC possessed a compact core–shell structure driven by strong protein–polysaccharide interactions, resulting in significantly enhanced thermal stability. SFN-ECC exhibited a significantly lower release rate during the simulated gastric phase. In conclusion, SFN-ECC created a structural barrier that efficiently protected SFN from degradation at acidic pH and gastrointestinal digestion while ensuring controlled intestinal delivery. This technology enables improving the bioavailability of unstable bioactive compounds, such as SFN, particularly for use in functional food formulations.

국문 초록 (Abstract)

설포라판(SFN)은 항산화 효능으로 잘 알려져 있으나, 구조적으로 불안정하여 위장관의 산성 조건에서 쉽게 분해된다. 따라서 SFN과 같은 지용성 화합물을 산성 위 환경에서 안정화하고 보호하기 위해서는 적절한 전달 시스템의 개발이 필요하다. 본 연구의 목적은 SFN의 안정성과 장 표적 전달을 향상시키기 위해, 복합 코아세르베이션을 통해 코팅된 SFN-적재 O/W 에멀젼(SFN-ECC)을 개발하는 것이었다. O/W 에멀젼은 수상에 Tween 80 (4 wt%)과 ι-카라기난을 용해하고, 유상에 해바라기유에 SFN을 용해하여 제조하였다. 이후 SFN-ECC 형성을 위해 녹두 분리 단백질(MPI)을 혼합하였다. 본 연구에서는 반응표면분석법을 두 단계에 걸쳐 적용하여 SFN-ECC를 최적화하였다. 1단계에서는 에멀젼의 주요 특성(안정성, 액적 크기, 분포)을 최적화하였으며, 2단계에서는 캡슐화 효율, 입자 크기, 코아세르베이트 수율에 초점을 맞추었다. 그 결과 도출된 최적 조건은 다음과 같다: 해바라기유 54 v/v%, ι-카라기난 0.60 w/v%, 8,793 rpm의 균질화 속도, pH 3.71, 그리고 MPI 1.17 w/v%. 최적화된 SFN-ECC는 강한 단백질–다당류 상호작용에 의해 형성된 조밀한 코어–셸 구조를 가지고 있었으며, 그 결과 열 안정성이 유의적으로 향상되었다. 또한 SFN-ECC는 모의 위 단계에서 유의적으로 낮은 방출률을 보이며 보호 효과를 입증하였다. 결론적으로 SFN-ECC는 산성 pH 및 소화 과정에서 SFN의 분해를 효과적으로 억제하는 구조적 장벽을 형성하면서, 장에서의 제어된 방출을 가능하게 한다. 본 기술은 SFN과 같이 불안정한 생리활성 물질의 생체 이용률을 향상시키기 위한 기능성 식품 제형에 유망한 접근법을 제공한다.

목차 (Table of Contents)

• 1. Introduction 1
• 2. Materials and methods 3
• 2.1. Materials 3
• 2.2. Optimization of SFN-loaded O/W emulsion formulation 3
• 2.2.1. Experimental design for the optimization of SFN-loaded O/W emulsion 3
• 2.2.2. Stability measurement of SFN-loaded O/W emulsion 4
• 2.2.3. Droplet size and distribution analysis 5
• 2.2.4. Multiple responses using desirability function 5
• 2.3. Optimization of SFN-E coated through complex coacervation formation 7
• 2.3.1. Experimental design for the optimization of SFN-loaded O/W emulsion coated through complex coacervation 7
• 2.3.2. Yield determination of SFN-loaded O/W emulsion coated through complex coacervation (SFN-ECC) 8
• 2.3.3. Particle size measurement 8
• 2.3.4. Determination of encapsulation efficiency (EE) 9
• 2.3.5. Multiple responses using desirability function 9
• 2.4. Fourier transform infrared spectroscopy (FTIR) 10
• 2.5. Confocal laser scanning microscopy (CLSM) 10
• 2.6. Thermogravimetric analysis (TGA) 11
• 2.7. In vitro digestion test 11
• 2.7.1. Simulated gastrointestinal digestion procedure 12
• 2.7.2. Sample preparation for LC-MS/MS analysis 12
• 2.7.3. Quantification of SFN by LC-MS/MS 13
• 2.7.4. Determination of cumulative SFN release rate 14
• 2.8. Statistical analysis 14
• 3. Results and discussion 16
• 3.1. Optimization of SFN-loaded O/W emulsion (SFN-E) using RSM 16
• 3.1.1. Effect of processing parameters on emulsion stability 19
• 3.1.2. Effect of processing parameters on droplet size 20
• 3.1.3. Effect of processing parameters on droplet distribution 22
• 3.1.4. Multiple response optimization of emulsion stability, droplet size, and distribution 23
• 3.2. Optimization of SFN-loaded O/W emulsion coated through complex coacervation (SFN-ECC) using RSM 26
• 3.2.1. Effect of pH and MPI concentration on coacervate yield 28
• 3.2.2. Effect of pH and MPI concentration on particle size 29
• 3.2.3. Effect of pH and MPI concentration on encapsulation efficiency 31
• 3.2.4. Multiple response optimization of coacervate yield, particle size, EE 33
• 3.3. FTIR analysis of sulforaphane-loaded structures 35
• 3.4. Visualization of the SFN-loaded O/W emulsion coated through complex coacervation (SFN-ECC) structure using CLSM 39
• 3.5. Thermal behavior of SFN-loaded O/W emulsion coated through complex coacervation (SFN-ECC) 41
• 3.6. SFN release behavior in a simulated gastrointestinal model 44
• 4. Conclusion 48
• References 49
• Appendix 58
• Abstract (in Korean) 66