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1 Vol.33 No.10 October 2015 Chinese Journal of Chromatography 1040 ~ 1045 DOI / SP.J / 1 1* / μm Carbohydrate 100 mm 2. 1 mm 2. 6 μm 0. 1% Target-MS / MS m / z ev - GF 2 GF 3 GF ppm 10 min GF 2 GF μg / kg GF 4 55 μg / kg mg / kg 3 GF 2 GF 3 GF % ~ % RSD 1. 6% ~ 8. 3% - / O658 A Determination of fructo-oligosaccharides in milk powder by high performance liquid chromatography- quadrupole / electrostatic field orbitrap high resolution mass spectrometry LIU Yun 1 DING Tao 1* XU Suli 2 WU Bin 1 SHEN Chongyu 1 ZHANG Rui 1 WANG Yan 1 FEI Xiaoqing 1 1. Animal Plant and Food Inspection Center Jiangsu Entry-Exit Inspection and Quarantine Bureau Nanjing China 2. School of Public Health Nanjing Medical University Nanjing China Abstract A method of high performance liquid chromatography-quadrupole / electrostatic field Orbitrap high resolution mass spectrometry HPLC-Q / Orbitrap MS was developed to deter- mine fructo-oligosaccharides in milk powder. The milk powder samples were dissolved in deion- ized water. Subsequently an aqueous solution of zinc acetate was used to precipitate protein. After centrifugation the final aqueous solution was filtered by a polytetrafluoroethylene PTFE membrane with pore size of μm. The analytes were separated on a Carbohydrate column 100 mm 2. 1 mm 2. 6 μm through gradient elution with the combination of acetoni- trile and 0. 1% formic acid aqueous solution. The target-ms / MS templates were performed at isolation window of m / z 4. 0 and collision energy of 30 ev in positive mode to extract the accu- rate product ion mass of analytes. Under the optimal condition 1-kestose GF 2 nystose *.Tel dingt@ jsciq.gov.cn. 2015KJ BAK19B BAK19B
2 10 - / 1041 GF 3 and 1-F-β-fructofuranosyl nystose GF 4 were well separated and the accuracy of extracted mass routinely detected was below ppm. The whole analysis time is only ten minutes. The detection limits for GF 2 and GF 3 were 100 μg / kg and the detection limit for GF 4 was 55 μg / kg. Good linearities were obtained in their respective linear ranges with correla- tion coefficients higher than The average recoveries at three spiked levels 5 10 and 20 mg / kg were in the range of 75. 8% % and the relative standard deviations RSDs were in the range of 1. 6% -8. 3%. The proposed method is simple sensitive fast and only in need of precipitation of proteins. The interference of matrix can be eliminated through the selection of product ion. The results were convenient and reliable and thus can be used in the large batch determination of any milk powder. fructo-oligosaccharides FOS - 11 HPLC-RID 12-1 ~ 3 β-1 2 HPLC-ELSD DP 2 ~ 9 16 HPLC-RID 1-kestose GF 2 nystose GF 3 1-F-β-fructofuranosyl nystose Key words high performance liquid chromatography-quadrupole / electrostatic field orbitrap mass spectrometry HPLC-Q / Orbitrap MS high resolution mass spectrometry HRMS fruc- to-oligosaccharides milk powder GF % ~ 2% HPLC- RID HPLC-ELSD HPLC-RID B HPLC-ELSD 9 GF 2 GF 3 GF 4 SPE 30 min inulo-triose F 2 inulo-tetraose F 3 inulo- - / pentaose F 4 9 AOAC GF 2 GF 3 GF 4 Q-Exac- tive HESI-Ⅱ Dionex 3000
3 XW-80A ms m / z 4. 0 Sigma LP % 99. 9% 99. 9% Merck TEDIA Milli-Q A 6% v / v B mg 0. 1 mg mmol / L C ph 5 mol / L ph 1. 90± min 10 ml 1. 0 g / L 5 mol / L ph 4. 70± S S mg / L μm 10 - Milk sample Precipitation Agilent Carbohydrate 100 mm 2. 1 mm 2. 6 μm 0. 1% A v / v acetic acid aqueous solution B 10 mmol / L zinc ace- B ml / min 10 μl tate solution C adjusting the ph based on the isoelectric 25 point. 0 ~ 2 min 80% B 2 ~ 5 min 80% B ~ % B 5 ~ 8. 5 min 20% B 8. 5 ~ 9 min 20% B ~ 80% B 9 ~ 10 min 80% B HESI-Ⅱ 320 N g 50 ml 20 ml 30 s 1 10 mmol / L ml 10 mmol / L 20 min r / min 5 min Table 1 Comparison of different precipitation methods for milk powder samples Recoveries / % GF 2 GF 3 GF 4 S1 A B C S2 A B C S1 domestic milk sample S2 foreign milk sample. A 6% Carbohydrate 100 mm L / min N 2 8 L / min N mm 2. 6 μm Carbohydrate 150 mm L / min 3 kv 50 V mm 5 μm Target-MS / MS R = ev AGC target 2e 5 100
4 10 - / a Target-SIM 2. 6 μm Target-MS / MS ev M+H + ESI + 2 GF 2 GF 3 GF 4 GF 2 GF 3 GF 4 M+H R b Fig. 1 1 GF 2 GF 3 GF 4 a b a First-full-scan accurate mass spectra and b second-extracted chromatograms of GF 2 GF 3 and GF 4 in milk samples Orbitrap Table GF 2 GF 3 GF 4 y x 3 S / N = 3 10 S / N = 2 Compound GF 2 GF 3 GF 4 Regression equations correlation coefficients r retention times t R limits of detection LODs n = 5 and limits of quantification LOQs n= 5 of the GF 2 GF 3 and GF 4 compounds Regression equation 10 2 GF 2 GF 3 GF 4 r r t R / min LOD / LOQ / μg / kg μg / kg GF 2 y = x GF 3 y = x GF 4 y = x y peak area x mass concentration mg / L. Linear range mg / L.
5 GF 2 GF 3 GF 4 Fig. 2 Second-full-scan accurate mass spectra of GF 2 GF 3 and GF ~ 2. 0 mg / L 5 GF 2 GF 3 GF S % ~ % RSD 1. 6% ~ 8. 3% S mg / kg 3 Table GF 2 GF 3 GF 4 n= 5 Recoveries of GF 2 GF 3 and GF 4 spiked in two different kinds of milk samples n= 5 Milk sample Spiked / mg / 100 g GF 2 Recovery / % RSD / % GF 3 Recovery / % RSD / % GF 4 Recovery / % RSD / % S S LC-MS / MS
6 10 - / 1045 Table n= 5 Determination results and labeled values of fructo-oligosaccharides FOS in ten milk powder samples n= 5 Sample No. GF 2 / mg / 100 g GF 3 / mg / 100 g GF 4 / mg / 100 g Total / mg / 100 g Labeled value / mg / 100 g GB Robefrorid M B. Am J Clin Nutr / Chromatography AOAC Official Method Medicine tography. 1 Zhang W. Journal of Biological Jiang B Zhang T Wang Z. Journal of Wuxi University for Light Industry. 15 Wang M H Li M Guo J et al. Chinese Journal of Health Inspection. 3 Chen M X Zhu W J. Biology Teaching Gnoth M J Kunz C Kinne-Saffran E et al. J Nutr Gao X J. Liquor Making Suppl Geng L J Huang J R Feng F et al. Chinese Journal of Wei F Wu Z Y Ma L Y et al. Asia Pacific Traditional. 12 Ding H L Li C Jin P et al. Chinese Journal of Chroma Liu Y F Li D Tang H C et al. Journal of Dairy Science and Technology. 17 Bai J Jiang J D Tao D L. Food Science Wang Y Z Yao P J Wei Y A. Food Industry Lin S J Xiao H L Guo B K. Chinese Journal of Health Inspection Zhang Y Y Nie S P Wan C et al. Food Science
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