Betulin, a pentacyclic triterpenoid, abundantly accumulated in Inonotus obliquus (chaga mushroom), exhibits strong anti-inflammatory, antioxidant, anticancer, and wound-healing properties. However, its extraction remains challenging due to its poor solubility and thermal sensitivity. In this study, we optimized ultrasound-assisted extraction (UAE) using response surface methodology (RSM) to maximize simultaneous betulin and antioxidant compound recovery from I. obliquus. We evaluated three extraction variables (i.e., time, temperature, and ethanol concentration) using a three-factor, five-level design. All quadratic models were significant (p < 0.05), with R² values ranging from 0.83 to 0.93 and prediction errors remaining below 5 %, thereby confirming strong model reliability. Multi-response optimization using a superimposed response plot identified 92.08 % ethanol, 42.56 min, and 62.19 °C as a narrow optimal region, in which all responses simultaneously met the desired criteria. Under these conditions, extraction was predicted to yield high phenolic content (2.58 mg GAE/g DM), increased flavonoid levels (0.57 mg QE/ g DM), strong DPPH radical-scavenging activity (87.49 % DSA), and a betulin content of 2.00 mg/g DM. In contrast, low ethanol concentrations, excessive heating, or prolonged extraction times resulted in reduced yields due to the oxidative or thermal degradation of the bioactive constituents. Overall, the optimized extraction conditions emphasize the importance of controlling solvent polarity and balancing the temperature and time parameters to prevent thermolabile compound decomposition. These results provide a reproducible and eco-efficient framework for large-scale antioxidant constituent extraction from I. obliquus.

ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
    재료 및 추출
    초음파 보조 추출공정
    총 폴리페놀 및 플라보노이드 함량 분석
    항산화 활성 평가
    베툴린 정량 분석
    반응표면분석법 설계
    회귀모델 구축 및 통계적 평가
RESULTS AND DISCUSSION
    모델 적합성 평가
    추출조건이 총 폴리페놀 함량(TPC)에 미치는 영향
    추출조건이 총 플라보노이드 함량(TFC)에 미치는 영향
    추출조건이 항산화 활성에 미치는 영향
    주요 생리활성 성분 분석
    추출조건이 베툴린 추출에 미치는 영향
    최적 추출조건 확인
CONCLUSION
REFERENCES

• 한민호(선문대학교 식품공학,영양학부) | Min Ho Han (Department of Food Technology & Nutrition, Sunmoon University 70, Sunmoon-ro 221 beon-gil, Tangjeong-myeon, Asan-si, Chungcheongnam-do, Korea)
• 오종현(선문대학교 식품공학,영양학부) | Jong Hyun Oh (Department of Food Technology & Nutrition, Sunmoon University 70, Sunmoon-ro 221 beon-gil, Tangjeong-myeon, Asan-si, Chungcheongnam-do, Korea)
• 이주환(선문대학교 식품공학,영양학부) | Ju Hwan Lee (Department of Food Technology & Nutrition, Sunmoon University 70, Sunmoon-ro 221 beon-gil, Tangjeong-myeon, Asan-si, Chungcheongnam-do, Korea)
• 김진우(선문대학교 식품공학,영양학부, 선문대학교 차세대 반도체 기술센터) | Jin Woo Kim (Department of Food Technology & Nutrition, Sunmoon University 70, Sunmoon-ro 221 beon-gil, Tangjeong-myeon, Asan-si, Chungcheongnam-do, Korea, Center for Next-Generation Semiconductor Technology, Sunmoon University, 70 Sunmoon-ro 221, Tangjeong-myeon, Asan-si, Chungnam 336-708, Korea) Corresponding author