돼지고기 중 5종의 곰팡이독소(아플라톡신 B1, B2, G1, G2 및 오크라톡신 A)를 분석하기 위해 0.1% 개미산을 함유한 50% 아세토니트릴용액으로 추출한 후 고체상추출칼럼을 이용하여 정제하고, LC-MS/MS로 동시 정량할 수 있는 시험법을 마련하였다. 분석조건으로 측정한 5종의 곰팡이독소 matrix-matched 표준곡선식에서 모두 상관계수 0.998이상의 상관관계를 나타내었다. 5종의 곰팡이독소 2배의 정량한계에서 10배의 정량한계로 첨가한 시료에서 평균 회수율은 72.1~109.9%로 실험 결과들이 EU 가이드라인에서 제시하는 유효성 확인을 위한 기준을 만족함으로써 시험법의 신뢰성을 확보할 수 있었다. 충청지역 유통되고 있은 돼지고기 20건에 대한 총아플라톡신 및 오크라톡신A에 대한 오염도 조사결과 정량한계 미만으로 조사되었다.

LC-MS/MS를 이용한 말라카이트 그린 유효성 검증결과 결정계수(R2)가 0.9996이상으로 높은 직선성을 보여주었으며 회수율은 어종에 따라 69.8%에서 93.9%범위를 보였다. 검출한계는 0.013 μg/kg, 정량한계는 0.038 μg/kg으로 CODEX의 가이드라인을 만족하였다. 수거된 어류 200건에서 말라카이트 그린을 분석한 결과, HPLC-DAD로는 1건이 검출되었으나, LC-MS/MS로 분석시 검출률은 18.5%였다. 본연구 결과, LC-MS/MS를 이용한 말라카이트 그린 분석법이 우리나라에서 많이 소비되는 담수어종에서 분석에 적합하며, 1건을 제외한 모든 시료에서 식품공전의 잔류허용기준 이하로 말라카이트 그린이 검출되어서 안전한 것으로 나타났다. 다만 식품공전의 검출한계 이하로 말라카이트 그린이 검출되는 빈도가 18.5%임을 볼 때, 지속적으로 감시하고 모니터링 할 필요가 있음을 시사한다.

이 연구는 축산물 중 포레이트 및 대사산물 5종의 안전관리를 위한 공정분석법을 확립하기 위하여 수행하였으며 분석법의 선택성, 검출한계 및 정량한계, 회수율에 대한 검증을 통하여 포레이트 및 대사산물 5종의 공정시험법으로의 유효성을 확인하였다. 포레이트 및 대사산물 5종을 신속하고 효과적으로 동시에 분석하기 위하여 LC-MS/MS를 사용하였고, 1% 아세트산 포함 아세토니트릴 추출 후PSA, C18을 이용해 정제하였다. 개발된 분석법의 평균 회수율은 79.2-113.9%였으며, 분석오차는 19.2% 이하로 정확성 및 재현성이 우수함을 확인할 수 있었다. 개발된 분석법은 국제적 잔류농약 분석 가이드라인에 적합한 수준이었으며 축산물 중 포레이트 및 대사산물 5종의 잔류검사를 위한 공정분석법으로 활용할 수 있을 것으로 판단된다.

농산물 중에 있는 제초제 saflufenacil의 잔류량을 검사 하기 위해 HPLC-UVD와 LC-MS를 이용한 정확하고 감도가 좋은 분석방법을 개발하였다. Saflufenacil 잔류물은 acetone 추출, dichloromethane을 이용한 액-액 분배, silica 와 carbon 카트리지 정제를 거쳐 기기분석을 수행하였다. 검량선 작성을 위해 0.1~5.0 μgmL−1 범위로 표준품을 만들어 실험한 결과 상관계수(r2)는 0.999로 높은 직선성을 보였다. 0.02~0.5 mg kg−1 처리수준으로 회수율을 실험한 결 과는 80.5~110.2% 이었으며, 상대표준편차는 10% 미만이었다. 분석방법의 검출한계와 정량한계는 각각 0.005와 0.02 mg L−1 이었다. 확립된 시험법으로 본청, 부산지방식품의 약품안전청과 경인지방식품의약품안전청에서 실험실간 검증을 실시한 결과 만족스런 결과를 얻었다. 이러한 결과 들을 통해 확립된 시험법은 농산물 중 saflufenacil의 잔류 량 분석에 적합함을 확인할 수 있었다.

The objective of this study was to develop a simultaneous method of 8 penicillin antibiotics including amoxicillin, ampicillin, cloxacillin, dicloxacillin, nafcillin, oxacillin, penicillin G and penicillin V in meat using LC-MS/MS. The procedure involves solid phase extraction with HLB cartridge and subsequent analysis by LC-MS/ MS. To optimize MS analytical condition of 8 compounds, each parameter was established by multiple reaction monitoring in positive ion mode. The chromatographic separation was achieved on a C18 column with a mobile phase of 0.05% formic acid and 0.05% formic acid in acetonitrile at a flow rate of 0.2 mL/min for 20 min with a gradient elution. The developed method was validated for specificity, linearity, accuracy and precision in beef, pork and chicken. The recoveries were 71.0~106%, and relative standard deviations (RSD) were 4.0~11.2%. The limit of detection (LOD) and the limit of quantification (LOQ) were 0.003~0.008 mg/kg and 0.01~0.03 mg/kg, respectively, that are below maximum residue limit (MRL) of the penicillins. This study also performed survey of residual penicillin antibiotics for 193 samples of beef, pork and chicken collected from 9 cities in Korea. Penicillins were not found in all the samples except a sample of pork which contained cloxacillin (concentration of 0.08 mg/kg) below the MRL (0.3 mg/kg).

A simultaneous determination was developed for 9 aminoglycoside antibiotics (amikacin, apramycin, dihydrostreptomycin, gentamicin, hygromycin B, kanamycin, neomycin, spectinomycin, and streptomycin) in meat by liquid chromatography tandem mass spectrometry (LC-MS/MS). Each parameter was established by multiple reaction monitoring in positive ion mode. The developed method was validated for specificity, linearity, accuracy, and precision based on CODEX validation guideline. Linearity was over 0.98 with calibration curves of the mixed standards. Recovery of 9 aminoglycosides ranged on 60.5~114% for beef, 60.1~112% for pork and 63.8~131% for chicken. The limit of detection (LOD) and limit of quantification (LOQ) were 0.001~0.009 mg/kg and 0.006~ 0.03 mg/kg, respectively in livestock products including beef, pork and chicken. This study also performed survey of residual aminoglycoside antibiotics for 193 samples of beef, pork and chicken collected from 9 cities in Korea. Aminoglycosides were not found in any of the samples.

Cotesia plutellae has been known as a natural enemy against the Diamondback moth, Plutella xylostella via laying eggs into a larva. When the larva hatches from the egg, teratocytes also are released and expected to work as immune suppressor via secreting immune suppressive factors such as venom proteins, teratocytes and polydnavirus. In order to identify immune suppressive factors from teratocytes, we collected the supernatant from serum-free culture media of teratocytes. Concentration of secreted proteins from teratocytes was successfully performed with using centricons among tested methods such proteins precipitation and electrophoresed in sodium dodecyl sulfate polyacrylamide gel. The gel slices were directly digested with trypsin using in-gel digestion method and analyzed via LC-Ms-Ms. Molecular weight of peptide fragments were compared with protein database predicted by full-genome sequences of C. plutellae. We identified two immune responsive proteins, which are calreticulin, host cellular response-related gene and neprilysin 2, immune regulator. This result suggests that host immune response is suppressed or regulated by the immune suppressive factors of teratocytes.

소염·진통 의약품 성분인 beclomethasone, dexamethasone, prednisolone, ketoprofen, phenylbutazone 5종에 대 한 동시분석을 위해 LC-MS/MS 분석 조건 중 MRM 방식 으로 각 성분의 MS 분석 최적조건을 결정하고, 정량이온 으로 beclomethasone 409.1/391.0, dexamethasone은 393.1/ 372.9, prednisolone은 361.0/343.1, ketoprofen은 255.0/209.0, phenylbutazone은 309.1/160.1 을 분석하였다. 혼합표준용 액을 기울기용매 조건으로 이동상 A(0.1% 개미산), B(0.1% 개미산을 함유한 아세토니트릴)를 이용하여 17분 동안 분 석한 결과, prednisolone (tR: 7.34 min), dexamethasone (tR: 7.73 min), beclomethasone (tR: 7.82 min), phenylbutazone (tR: 8.31 min), ketoprofen (tR: 9.24 min) 순서로 검출되었 다. 5종 성분 모두 약 2-100 ng/mL 농도 수준의 검정곡선 에서 R2 값이 0.999 이상의 우수한 직선성을 나타내었고, prednisolone을 제외한 4종 성분의 검출한계는 0.4-0.9 ng/ mL, 정량한계는 0.81-2.22 ng/mL 였으며, prednisolone은 그보다 다소 높은 4.60, 11.46 ng/mL 이었다. 환제품의 기 타가공품 공시료에 5종 성분 혼합표준용액을 최종농도가 5, 20, 50 ng/mL 이 되도록 직접 첨가하여 분석한 결과, 모든 농도에서 80% 이상의 우수한 회수율을 얻을 수 있 었고, 일내 일간 반복 분석의 상대표준편차(%)를 통해 모 든 성분에서 비교적 재현성 있는 결과가 나타났다. 실제 인터넷 상에서 유통중인 식품에 이 분석법을 적용한 결과 모든 제품에서 검출되진 않았으나, 소비자들의 건강상 위 해를 끼칠 수 있는 이들 성분의 동시분석법을 활용한 지 속적인 모니터링이 필요한 실정이다.

감귤류의 껍질은 플라보노이드의 중요한 소스 중의 하나로서 동아시아에서 내장 및 염증성 질환을 치료하는 민간 의약품으로서 사용되어 왔다. 본 연구에서는 하귤 (C. natsudaidai) 껍질에 포함되어 있는 플라보노이드 성분을 고성능액체크로마토그래피-이중질량분석법 (HPLC-MS/MS)으로 10 개성분을 동정하였다. 플라바논, 플라본 및 쿠마린 유도체는 각각 hesperetin, noviletin 및 coumarin을 사용하여 유효화 하였으며 유효화된 방법으로 정량하였다. 상관계수 (r2)는 > 0.9970으로서 높은 값을 보여주었다. LOD는 >0.01 mg/L이었으며 LOQ는 >0.05 mg/L이었다. 플라보노이드의 총량은 9229.7 ± 0.5 mg/kg 이었다. Naringin의 량이 5010.0 ± 4.5 mg/kg으로 가장 많았으며 sinensetin의 량이 0.6 ± 0.1 mg/kg으로 가장 적었다. 항산화력을 25 μg/mL 에서 500 μg/mL의 농도범위에서 DPPH·, ABTS·+, NO· 소거능 및 FRAP의 항으로 분석하였다. 감귤 플라보노이드의 항산화 능력은 시료의 량이 증가하면 증가함을 알 수 있다.

A rapid, simple and reliable LC-MS/MS method, which can be used on a routine basis, was developed for the simultaneous detection of 8 penicillin antibiotics (amoxicillin, ampicillin, penicillin-G, penicillin-V, oxacillin, cloxacillin, nafcillin and dicloxacillin) in swine muscle and kidney. The antibiotics were extracted from samples with water and methanol. The extract was centrifuged, filtered and analyzed by liquid chromatography coupled with a tandem mass spectrometer (LC-MS/MS), using a C18 reversed phase column with water/acetonitrile gradient containing 0.05 % formic acid. Mass spectral acquisition was achieved in an electrospray positive ion mode by applying multiple reaction monitoring (MRM) of 2 fragment ion transitions to provide a high degree of sensitivity and specificity. The ion rations were consistent and could be used for confirmation of identity of the penicillin antibiotics. Recoveries of eight penicillins at three fortification levels (10, 20 and 40㎍/㎏) ranged from 79.8 to 102.4% and 72.8 to 103.4% in swine muscle and kidney, respectively. The coefficient of variation was than 9% in all samples. The estimated limits of quantification (LOQs) ranged from 1.0 to 3.2㎍/㎏ in swine muscle and kidney, respectively. The LOQs of this method are below the MRLs of penicillin antibiotics in animal tissues established in Korea and other countries.

A reversed-phase high-performance liquid chromatography with tandem mass-spectrometric detection was developed and validated for the simultaneous analysis of 13 quinolones (orbifloxacin, sarafloxacin, marbofloxacin, ofloxacin, enrofloxacin, danofloxacin, pefloxacin, ciprofloxacin, norfloxacin, difloxacin, oxolinic acid, flumequine, nalidixic acid) in pork. The drugs were extracted from homogenized sample using acidic buffer (2% trichloracetic acid) and acetonitrile with ethyl acetate. Analysis was performed by liquid chromatograph with tandem mass spectrometry (LC-MS/MS) in positive mode for all quinolones. Mass spectral acquisition was performed in the multi reaction monitoring mode (MRM), selecting two structurally significant transitions per compound. Good linear relationship (R2=0.999) was observed at 6 concentrations of 2.5～100 ng/g. Satisfied recoveries (73.3～105.4%) of all quinolones were demonstrated in spiked pork at three levels from 5 to 20 ng/g. The limits of quantitation (LOQs) for quinolones ranged from 0.25 to 5.0 ng/g.

Metabolomics aims at the comprehensive, qualitative and quantitative analysis of wide arrays of endogenous metabolites in biological samples. It has shown particular promise in the area of toxicology and drug development, functional genomics, system biology and clinical diagnosis. In this study, analytical technique of MS instrument with high resolution mass measurement, such as time-of-flight (TOF) was validated for the purpose of investigation of amino acids, sugars and fatty acids. Rat urine and serum samples were extracted by selected each solvent (50% acetonitrile, 100% acetonitrile, acetone, methanol, water, ether) extraction method. We determined the optimized liquid chromatography/time-of-flight mass spectrometry (LC/TOFMS) system and selected appropriated columns, mobile phases, fragment energy and collision energy, which could search 17 metabolites. The spectral data collected from LC/TOFMS were tested by ANOVA. Obtained with the use of LC/TOFMS technique, our results indicated that (1) MS and MS/MS parameters were optimized and most abundant product ion of each metabolite were selected to be monitorized; (2) with design of experiment analysis, methanol yielded the optimal extraction efficiency. Therefore, the results of this study are expected to be useful in the endogenous metabolite fields according to validated SOP for endogenous amino acids, sugars and fatty acids.

LC-DAD-ESI/MS를 이용하여 국내 자색벼 품종에 대해 개별 안토시아닌 조성 및 함량을 평가한 결과는 다음과 같다. 1. 자색벼 품종에서 분리된 모든 개별 안토시아닌의 화학구조는 MS fragment 패턴을 확인하여 cyanidin을 base로 한 unknown 화합물 1종을 포함, cyanidin 3,5-diglucoside, cyanidin 3-glucoside, peonidin 3-glucoside의 총 4가지 개별성분이 분리 및 동정되었다. 2. Cyanidin 3-glucoside 및 peonidin 3-glucoside이 주요성분으로 cyanidin 3-glucoside의 경우 90% 이상의 가장 높은 함량 비중을 나타냈으며, 개별성분별 평균 함량은 cyanidin 3-glucoside > peonidin 3-glucoside > cyanidin 3,5-diglucoside > unknown(cyanidin based)의 순으로 나타났다. 3. 흑진주벼의 총 안토시아닌 함량은 408 mg/100 g으로 흑남벼보다 약 6배 정도 높은 함량을 나타내었다.

A simple, selective and sensitive procedure for the confirmation of 14 sulfonamide antibacterials in milk was developed. The milk samples were homogenized, extracted and deproteinized by acetonitrile and defatted by n-hexane. Analysis was performed by liquid chromatography with tandem mass spectrometry (LC-MS/MS) in positive mode for all 14 analytes. Mass spectral acquisition was performed in the multi reaction monitoring mode (MRM), selecting two structurally significant transitions per compound. The calibrations were performed in sample matrixes and the interference effect of sample matrixes on the ionization was effectively eliminated. Good linear relationship (R2=0.992～ 0.999) were observed at 6 concentrations of 2.5～100 ng/g. Satisfied recoveries (86.3～110.2%) of all sulfonamides were demonstrated in spiked milk at three levels from 5 to 20 ng/g. The limits of quantitation (LOQs) for sulfonamides ranged from 0.25～2.1 ng/g.

The pomegranate (Punica granatum), especially its fruit, possesses a vast ethnomedical history and represents a phytochemical reservoir of heuristic medical value. The tree and fruit can be divided into several anatomical compartments, and the fruit juice, peel and oil are known to be weakly estrogenic and heuristically of interest for treatment of menopausal symptoms and sequellae. In this study, analysis of estrogen in pomegranate extract was carried out with LC/MS/MS. Three batches of pomegranate extract samples were used to analysis the target compounds (estrogen). The contents of estrogen derivatives in the samples were 38.6 ppb of estriol, 83.5 ppb of estrone,and 10.9 ppb of estradiol. This result suggests that the pomegranate extract can used for treatment of menopause symptoms in the woman.

In order to cope with risk issues of overseas on 4,4'-diaminodiphenylmethane (4,4'-MDA) known to used in the production of polyamide resins, we investigated on 4,4'-MDA migration from plastic cooking utensils. 93 samples of plastic cooking utensils were collected from the retail markets in 7 major cities of Korea during March to June, 2007 and 4,4'-MDA levels migrated to food simulants, water, 4% acetic acid, 20% ethanol and n-heptane, on plastic cooking utensils was quantified by liquid chromatography with tandem mass spectrometry (LC-MS/MS). 4,4'- MDA was detected in 14 samples, and the levels of 4,4'-MDA migrated to food simulants were in the range of 0.001 ~ 0.026 ppm. Among them, the 4,4'-MDA migration amounts of 2 samples were above 0.01 ppm defined by the EU.