초록 ▼

○ 연구목표
본 연구의 목표는 간장의 소비가 많은 우리나라를 고려하여 간장의 유해 독성물질인 3-MCPD의 표준화된 분석방법을 확립하여 극미량 및 신뢰성이 보장되는 표준분석기법을 개발·확립하고자 하였다.
○ 연구내용
3-MCPD 분석을 위한 표준화된 분석법 확립을 위해 ① 유도체 반응조건 선정 ② 다양한 흡착제에 대한 방법 시도 ③ 효과적인 용출용매 선정 ④ Surrogate, 내부표준물질(internal standard) 물질 선정 ⑤ 3-MCPD의 표준화된 분석법 검증의 단계로 진행함.
○ 연구개발결과

○ 연구목표
본 연구의 목표는 간장의 소비가 많은 우리나라를 고려하여 간장의 유해 독성물질인 3-MCPD의 표준화된 분석방법을 확립하여 극미량 및 신뢰성이 보장되는 표준분석기법을 개발·확립하고자 하였다.
○ 연구내용
3-MCPD 분석을 위한 표준화된 분석법 확립을 위해 ① 유도체 반응조건 선정 ② 다양한 흡착제에 대한 방법 시도 ③ 효과적인 용출용매 선정 ④ Surrogate, 내부표준물질(internal standard) 물질 선정 ⑤ 3-MCPD의 표준화된 분석법 검증의 단계로 진행함.
○ 연구개발결과
① 각 유도체의 머무름 시간은 HFB-$d_5$-MCPD 및 HFB-MCPD가 각각 5.71 및 5.77min이었으며, PB-$d_5$-MCPD 및 PB-MCPD은 12.33 및 12.37min이였음. 또한 TFA를 이용한 TMS-$d_5$-MCPD 및 TMS-MCPD의 머무름시간은 각각 8.10 및 8.14min이였으며, MBTFA를 사용한 TFA-$d_5$-MCPD 및 TFA-MCPD 유도체방법의 머무름시간은 각각 3.98 및 4.01min으로 관찰됨.
② HFBI, PBA 및 BSTFA를 사용한 유도체 3가지 방법 모두 10ppb이하의 검출한계를 나타냄으로써 현재의 국내 0.3ppm기준을 검사해내는데는 무리가 없는 것으로 평가됨.
③ PBA 및 BSTFA 유도체방법은 후처리가 필요없어 손쉬운 처리가 가능하였으며 특히 PBA 유도체화 방법은 가열이 필요없어 가장 짧은 시간에 반응이 가능하였음.
- HFBI에 의한 acylation방법은 ECD를 사용할 경우 검출한계가 2∼3ppb수준까지도 낮출 수 있어 가장 민감한 방법으로 평가되지만 30분 반응 후 물로 닦아내는 과정을 거쳐야 하는 번거로움이 단점.
- Extrelut20을 대체할 수 있는 adsorbent로는 florisil이 가능한 것으로 나타났음.
- 국내 공전에 제시되어 있는 ethyl acetate와 AOAC 등재방법에서 사용하는 diethyl ether의 추출능을 살펴본 결과, ethyl acetate가 보다 효과적으로 간장으로부터 3-MCPD를 회수해 내었으며 hexane등에 의한 비극성 방해물질제거는 효과가 없었음.
- Surrogate 및 internal standard는 머무름시간 및 검출기의 감도등을 고려하여 약 5종의 대상성분 중 각각 1,2-butanediol 및 1,2-dibromo-3-chloropropane을 최종 선정하였음.
○ 연구성과(응용분야 및 활용범위포함)
본 연구의 성과는 식품 중 3-MCPD(3-monochloropropane-1,2-diol)의 효율적인 분석법을 제시함으로써 국내 섭취량이 빈번한 간장 중 발암가능물질인 3-MCPD의 규제 방안을 확립할 수 있는 기틀을 마련하였고 본 연구에서 확인된 시험방법의 국제화를 통해 국내의 공인된 분석법이 세계적인 표준기법으로 제시될 수 있을 것임.

Abstract ▼

The contamination levels of 3-monochloro-1,2-propanediol(3-MCPD) and 1,3-dichloro- propanol(1,3-DCP) in soy sauce, sauces, HVP and Ramyun soup base were monitored. 442 samples were collected from April 2002 to October 2002 in Seoul, Kyung-ki, Kyung-sang, Choong-chung, Kang-won and Chun-la. 3-MCPD wa

The contamination levels of 3-monochloro-1,2-propanediol(3-MCPD) and 1,3-dichloro- propanol(1,3-DCP) in soy sauce, sauces, HVP and Ramyun soup base were monitored. 442 samples were collected from April 2002 to October 2002 in Seoul, Kyung-ki, Kyung-sang, Choong-chung, Kang-won and Chun-la. 3-MCPD was analyzed by GC-MSD. 1,3-DCP was analyzed by GC-ECD. As results, the detection range of 3-MCPD concentration was between 0.02㎎/㎏ and 0.82㎎/㎏ in soy sauce, 0.04㎎/㎏∼0.48㎎/㎏ in sauces, 0.03㎎/㎏∼0.28㎎/㎏ in HVP, 0.02㎎/㎏∼0.14㎎/㎏ in Ramyun soup base respectively. However any samples were not detected with 1,3-DCP. The contaminated rates of 3-MCPD in soy sauce was 36.9%, sauce was 5.8%, HVP was 16.7% and Ramyun soup base was 44.1%.
Soy sauce is the representative vegetable protein hydrolyzed food. The reliable analytical method for the toxic 3-MCPD in Soy sauce has been studied.
Following 5 steps were proceeded to develope the analysis method: i) Optimization of derivatization methods ii) Searching for effective adsorbents iii) Selection of good elution solvents iv) Selection of surrogate and internal standard and v) Validation of total analytical methods.
The derivatization studies of 3-MCPD were performed mainly as acylation with HFBI(Heptafluorobutyrylimidazole), alkylation with PBA(Phenylboric acid) and silylation with MBTFA(N-Methyl-bis[trifluoroacetamide]). Other derivatization methods like tert.-Butyl-dimethylsilylation with MTBSTFA and acylation with MBTFA (N-Methyl-bis [trifluoroacetamide]) were also considered.
The retention times of derivative, HFB-$d_5$-MCPD, HFB-MCPD, PB-$d_5$-MCPD and PB-MCPD, were 5.71, 5.77, 12.33 and 12.37 minutes, respectively and TMS-$d_5$-MCPD and TMS-MCPD were eluted at 8.10 and 8.14 minutes, respectively. Retention time of TFA-$d_5$-MCPD and TFA-MCPD derivatized with MBTFA, was 3.98 and 4.01 minutes, respectively which are eluted too early for GC-MSD analysis. The TBDMS derivative of 3-MCPD was not eluted at all. The derivatives of 3-MCPD with HFBI, PBA and BSTFA showed below 10ppb which was sensitive enough to satisfy Korea maximum residue limit 0.3 ppm. In respect to convinience of analytical method, PBA-derivatizing method was the most simple due to no heating was needed. Silylation with BSTFA and acylation with HFBI were needed to heat 60 and 30 minutes, respectively. The acylation with HFBI was cumbersome due to washing of the excess HFBI reagent. The acylation method with HFBI was the most sensitive method showing the lowest detection limit, 2∼3 ppb with electron capture detector.
The adsorbents to purify soy sauce for 3-MCPD with matrix dispersion method were screened with Celite (Sigma), Silicagel (Merck), Florisil (J. T. Baker) and LC-Diol (Alltech) with Extrelut20 (Merck). Florisil was turned out the unique alternative adsorbent being able to be used for the purification of soy sauce for 3-MCPD instead of Extrelut20. Others were not sufficient to recover 3-MCPD enoughly. Ethyl acetate was most efficient to elute 3-MCPD with good recovery rate better than diethyl ether which was suggested by AOAC published method. As the experiment to clean-up of endogenous nonpolar interferences in soy sauce samples, washing the adsorbent with hexane-containing solvent before ethyl acetate or diethyl ether elution was not effective.
The internal standard heptadecane recommended in Korea Food Code might be intrinsic compound in soy sauce and the deuterilated 3-MCPD for isotope dilution method was too expensive to be used in common laboratories. Therefore the replacement for them were screened. Internal standard was considered for good sensitivity and retention time on instrument and the good recovery rate through total analysis was searched for surrogate compound. 1,2-butanediol and 1,2-dibromo-3-chloropropane were selected as the surrogate and internal standard among more than five chemicals.
The validation study for the above three derivatization methods were performed to check the recovery, reproducibility and sensitivity for 3-MCPB spiked soy sauce sample.
The average recovery of three method was 92.25% and the average percent relative standard deviation was 1.045% which are quite sufficient to be used for 3-MCPD analysis in soy sauce. The limit of detection for PB-MCPD and TMS-MCPD were 10.16 and 7.06ppb on GC/MSD, respectively. HFB-MCPD derivative showed the lowest detection limit 2.98ppb on GC/ECD and the limit of detection on GC/MSD was 5.32ppb.

목차 Contents

• 표지 ...1
• 제출문 ...2
• 요약문 ...4
• 영문요약문 ...10
• 목차 ...12
• I. 서 론...14
• II. 국내.외 기술개발 현황...16
• III. 제1세부연구개발과제 결과...18
• 제1절 재료 및 방법...20
• 1. 재료와 시약 및 3-MCPB의 분석...20
• 2. 재 료와 시 약 및 1,3-DCP의 분석...29
• 제2절 결과 및 고찰...33
• 1. 시중에 판매되고 있는 간장, 소스류, HVP, 복합조미식품 (스프류)의 3-MCPB 오염정도 및 실험결과...33
• 2. 3-MCPD가 검출된 시료에서의 1,3-DCP 오염정도...37
• 3. 식푼중의 B-MCPB 오염도에 관한 고찰...38
• 제3절 연구개발목표 달성도 및 대외 기여도...39
• 제4절 연구개발결과의 활용성과 및 계획...40
• 제5절 기타 중요변경사항...41
• 제6절 참고문헌...42
• 제7절 연구과제(세부과제)요약서...44
• IV. 제2세부연구개발과제 결과...58
• 제1절 연구개발수행 내용...60
• 제2절 연구개발수행 과정...61
• 1. 식품중 3-MCPD의 분석동향 및 분석방법에 관한 최신정보수집 및 정리
• 2. 기존 분석방법 결과와 새로 개발된 분석방법의 비교...61
• 3. MCPD 분석을 위한 표준화된 분석법 확립...68
• 제3절 연구개발수행 결과...87
• 1. 3-MCPD 분석을 위한 표준화된 분석법 확립...87
• 가. 유도체 반응 조건 선정...87
• 나. 대체 흡착제 선정에 관한 연구...109
• 다. 효과적인 용출용매 선정...111
• 라. Surrogate, 내부표준물질(internal standard) 선정...114
• 마. 3-MCPD 분석을 위한 세가지 표준화된 분석법 검증...119
• 2. 3-MCPD 분석을 위한 분석법의 제안...121
• 제4절 연구개발목표 달성도 및 대외기여도...124
• 제5절 연구개발결과의 활용성과 및 계획...125
• 제6절 기타중요변경 사항...126
• 제7절 참고문헌...127
• 제8절 연구과제(세부과제)요약서...130