S Y BAN NHÂN DÂN TP.HCM KHOA H C VÀ CÔNG NGH BÁO CÁO NGHI M THU (Đã ch nh s a theo góp ý c a H i đ ng nghi m thu) NGHIÊN C U PHƯƠNG PHÁP PHÂN TÍCH CÁC CH T KÍCH THÍCH TĂNG TRƯ NG H BETAAGONIST(CLENBUTEROL, SALBUTAMOL) TRONG TH T HEO, GAN HEO, TH C ĂN NI HEO B NG PHƯƠNG PHÁP PHÂN TÍCH S C KÝ KHÍ GHÉP KH I PH CH NHI M Đ TÀI CƠ QUAN QU N LÝ CƠ QUAN CH QU N THÀNH PH H CHÍ MINH THÁNG 2/ 2009 S KHOA H C VÀ CƠNG NGH TP.H CHÍ MINH TRUNG TÂM D CH V PHÂN TÍCH VÀ THÍ NGHI M BÁO CÁO NGHI M THU NGHIÊN C U PHƯƠNG PHÁP PHÂN TÍCH CÁC CH T KÍCH THÍCH TĂNG TRƯ NG H BETA-AGONIST (CLENBUTEROL, SALBUTAMOL) TRONG TH T HEO, GAN HEO, TH C ĂN NUÔI HEO B NG PHƯƠNG PHÁP PHÂN TÍCH S C KÝ KHÍ GHÉP KH I PH GC/MSn CH NHI M Đ TÀI: THS CHU VÂN H I THÀNH PH H CHÍ MINH THÁNG 01/2009 S KHOA H C VÀ CƠNG NGH TP.H CHÍ MINH TRUNG TÂM D CH V PHÂN TÍCH VÀ THÍ NGHI M BÁO CÁO NGHI M THU NGHIÊN C U PHƯƠNG PHÁP PHÂN TÍCH CÁC CH T KÍCH THÍCH TĂNG TRƯ NG H BETA-AGONIST (CLENBUTEROL, SALBUTAMOL) TRONG TH T HEO, GAN HEO, TH C ĂN NI HEO B NG PHƯƠNG PHÁP PHÂN TÍCH S C KÝ KHÍ GHÉP KH I PH GC/MSn CH NHI M Đ TÀI: THS CHU VÂN H I THÀNH PH H CHÍ MINH THÁNG 01/2009 TĨM T T N I DUNG NGHIÊN C U Clenbuterol (CLEN), Salbutamol (SAL) h p ch t thu c nhóm beta-agonist đư c s d ng b t h p pháp th c ăn gia súc đ kích thích tăng trư ng, gi m hàm lư ng béo tăng hàm lư ng th t T i Vi t Nam, CLEN SAL đư c phân tích ch y u b i phương pháp sàng l c Elisa (Enzyme-Linked Immunosorbent Assay) Phương pháp GC/MS hay LC/MS chưa có phương pháp chu n nh t khâu chu n b m u T i Trung tâm d ch v phân tích thí nghi m Thành ph H Chí Minh (TTDVPTTN) nghiên c u phương pháp phân tích CLEN, SAL b ng GC/MS đư c s d ng đ phân tích CLEN, SAL th c ăn gia súc, th t heo gan heo, th c ph m ch bi n s n t i thành ph H Chí Minh t nh l n c n M u đư c ly trích b i dung d ch đ m CH3COONH4, enzym β_Glucuronidase sau đư c làm s ch b i c t SCX (Alltech SPE 500mg/6ml), r a gi i b ng 6ml MeOH:NH3 (95:5) Sau đư c d n xu t v i MSTFA + NH4I + DTE (1000:4:2) 600C 15 phút CLEN, SAL đư c phân tích b i GC/MS (ESI (+) SIM) v i m/z 227, 300, 335, 336 337 cho CLEN, 207, 294, 350, 369 370 cho SAL Hi u su t thu h i c a phương pháp 65%-88% Gi i h n phát hi n c a phương pháp cho CLEN 0,16 ppb cho th c ăn gia súc, 0,05ppb cho th t heo, 0,05ppb cho gan heo; cho SAL 0,11ppb cho th c ăn gia súc, 0,07ppb cho th t heo, 0,07ppb cho gan heo Phương pháp Elisa đư c s d ng đ phân tích m u th c ăn cho heo, th t heo, gan heo đư c xác nh n l i b ng GC/MS cho k t qu âm tính dương tính, có so sánh k t qu v i LC/MS đ i v i m u dương tính SUMMARY OF RESEARCH CONTENT Clenbuterol (CLEN), Salbutamol (SAL) may be illegally used as feed additive for growth promotion with concomitant reduction of fat tissues in meat-producing livestock In Viet Nam, CLEN, SAL are being mainly detected by the screening ELISA method Use of GC/MS (Gas chromatographic-mass spectrometric method) or LC/MS (Liquid chromatographic-mass spectrometric method) is somewhat limited due to lack of appropriate sample preparation Center of analytical services and experimentation (CASE) has devised a quite efficient GC/MS method for CLEN, SAL analysis which are being used to control feedstuff, pork meat, liver, processed food in Ho Chi Minh City and other provinces Sample preparation involves extraction of CLEN, SAL by a CH3COONH4, enzyme β_Glucuronidase followed by a clean up step with SCX (Alltech SPE 500mg/6ml) column and elution with 6ml MeOH:NH3 (95:5) The residue is derivatized with MSTFA + NH4I + DTE (1000:4:2) at 600C for 15 minutes The selected ions monitoring mode is performed at m/z 227, 300, 335, 336 and 337 for CLEN, and 207, 294, 350, 369 and 370 for SAL Recoveries of CLEN, SAL 65% 88% The limits of detection were about 0.16ppb_feedstuff, 0.05ppb_pork meat, 0.05ppb_ liver for CLEN; 0.11ppb_feedstuff, 0.07 ppb_ pork meat, 0.07ppb_ liver for SAL The technique include Elisa that is used as screening methods and GC/MS as confirmatory methods and is compared with LC/MS for positive samples M CL C Trang Tóm t t đ tài ti ng Vi t Tóm t t đ tài ti ng Anh M cl c Danh sách hình CHƯƠNG 1: M Đ U Tên đ tài 1 Ch nhi m đ tài Cơ quan ch trì Th i gian th c hi n Kinh phí đư c t Kinh phí c p M đ u 1.1.Tình hình c m s d ng ch t kích thích tăng trư ng th gi i Vi t Nam 1.2 Tình hình nghiên c u phân tích CLEN, SAL nư c 1.3 M c tiêu cu đ tài 1.4 Đ i tư ng nghiên c u 1.5 N i dung th c hi n đ tài 1.6 Phương pháp nghiên c u 1.7 Nơi thu th p m u 10 1.8 Th i gian thu th p m u 10 1.9 Nơi phân tích 10 1.10 S n ph m cu đ tài 10 1.11 Ý nghiã th c ti n 10 CHƯƠNG II: T NG QUAN 12 2.1 T ng quan v CLEN SAL 12 2.2 T ng quan phương pháp phân tích CLEN, SAL th c ăn 16 nuôi heo, th t heo, gan heo b ng GC/MS đăng báo 2.3 Sơ lư c v k thu t s c ký khí 18 2.4 Phương pháp s c ký khí ghép kh i ph 27 CHƯƠNG III: NGHIÊN C U PHƯƠNG PHÁP PHÂN TÍCH CHLEN, SAL TRONG TH T HEO, GAN HEO, TH C ĂN NUÔI HEO B NG K THU T GC/MS 39 3.1 Thi t b - Hoá ch t 39 3.2 Pha dung d ch 41 3.3 Kh o sát t i ưu ph n ng d n xu t sylyl hoá CLEN, SAL n i 41 chu n 17- Methyltestosteron chu n b ng GC/MS CHƯƠNG IV: KH O SÁT PHÂN TÍCH TRÊN M U TH T 58 4.1 X lý m u 58 4.2 Xác đ nh LOD máy 65 4.3 Đư ng chu n 66 4.4 Xác đ nh MLOD, MLOQ, hi u su t thu h i, đ ch m n n m u 68 th c ăn gia súc 4.5 Xác đ nh MLOD, MLOQ, hi u su t thu h i, đ ch m n n m u 71 th t heo 4.6 Xác đ nh MLOD, MLOQ, hi u su t thu h i, đ ch m n n m u 73 gan heo 4.7 Tính tốn đ khơng đ m b o cu m u 76 CHƯƠNG V: K T QU VÀ TH O LU N 78 5.1 Tiêu chí l a ch n đi m thu th p m u 79 5.2 Thu th p m u th c ăn, m u th t heo, gan heo, th c ph m ch bi n s n 79 5.3 K t qu th o lu n_ K t qu phân tích CLEN, SAL b ng Elisa 83 5.4 K t qu phân tích CLEN, SAL b ng GC/MS 86 5.5 Ki m soát ch t lư ng k t qu th nghi m 89 5.6 K t qu phân tích CLEN, SAL n n th c ph m ch bi n s n 92 CHƯƠNG VI: K T LU N VÀ Đ NGH 94 S N PH M C A Đ TÀI: Qui trình phân tích Clenbuterol, Salbutamol th t heo, gan heo, th c ăn cho heo PH L C TÀI LI U THAM KH O DANH SÁCH CÁC HÌNH TÊN HÌNH NH S TRANG 2.1 Sơ đ máy s c ký khí 18 2.2 Mặt cắt cột sắc ký mao quản phóng to 19 2.3 Mức độ phân cực loại cột sắc ký khí Alltech 20 2.4 Cấu trúc cột sắc ký 21 2.5 Thời gian lưu cấu tử phân tích 22 2.6 Sơ đồ loại sắc ký khí ghép khối phổ 27 2.7 Sự phân mảnh với thay đổi lượng bắn phá 30 2.8 Sơ đồ trình hoạt động máy sắc ký khí ghép khối phổ 30 2.9 Sơ đồ GC/MS loại b y ion 34 2.10 Sơ đồ Ion trap electrode 34 3.1 K t qu hi u ch nh máy (tuning) 43 3.2 S c ký đ DFBZ 10 ng/ml 44 3.3 Kh i ph c a d n xu t di-TMS-CLEN di-TMS-IS 47 3.4 Ph n ng t o d n xu t silyl hoá CLEN IS 48 3.5 Cơ ch phân m nh di-TMS-CLEN full MS 49 3.6 Cơ ch phân m nh di-TMS-(17-methyltestosterone) full MS 50 ch đ full MS ( EI ) CHƯƠNG 1: M Đ U Tên đ tài: Nghiên c u phương pháp phân tích ch t kích thích tăng trư ng h betaagonist (Clenbuterol, Salbutamol) th t heo, gan heo, th c ăn nuôi heo b ng phương pháp phân tích s c ký khí ghép kh i ph GC/MS Ch nhi m đ tài: Ths.Chu Vân H i Cơ quan ch trì: Trung tâm d ch v phân tích thí nghi m TPHCM Th i gian th c hi n : 12/2005-12/2008 Kinh phí đư c t : 210.000.000 VNĐ M đ u : Clenbuterol (CLEN) Salbutamol (SAL) ch t b B Nông nghi p phát tri n nông thôn (PTNT) c m s d ng t năm 2002 M c dù b c m ch t v n đư c lút s d ng th c ăn chăn nuôi S t n dư c a ch t gây nh ng r i l an ch c tim ph i tim đ p nhanh, run cơ, đau đ u, bu n nôn…mà ch t l i ch b phân gi i nhi t đ cao Hi n chưa có tài li u khoa h c nói ch t Clenbuterol Salbutamol dư lư ng c a chúng s n ph m chăn nuôi gây ung thư cho ngư i Tuy nh hư ng c a dư lư ng ch t đư c coi đ c trung bình nhà khoa h c kh ng đ nh r ng t t c nh hư ng x u c a ch t ngư i không th ch p nh n đư c Th i gian qua, m t s quan báo chí Đài truy n hình Vi t Nam đăng t i v vi c s d ng hócmơn kích thích sinh trư ng th c ăn chăn nuôi vi c qu n lý nhà nư c v lĩnh v c B Nơng nghi p PTNT có thông báo ý ki n ch đ o ngày 09/06/2006 s 2646/TB-BNN-VP v vi c ki m tra ch t lư ng Clenbuterol th c ăn chăn nuôi ph m vi tòan qu c Đ ng th i, ngày 25/08/2006 B trư ng b Nông nghi p PTNT ban hành ch th v vi c tăng cư ng công tác qu n lý, nâng cao ch t lư ng nông lâm s n, v t tư nông nghi p đ m b o v sinh, an tịan nơng s n, th c ph m Theo đó, B trư ng ch đ o đơn v tr c thu c thư ng xuyên ki m tra v ch t lư ng s n ph n chăn nuôi, gi ng th c ăn chăn nuôi B trư ng b Nông nghi p PTNT quy t đ nh ngày 09 tháng 01 năm 2007 s 65/QĐ-BNN-CN v MLOD M U TH C ĂN NL: 5.70E3 RT : 8.48 100 90 85 90 85 90 85 80 75 65 60 55 50 75 70 Relative Abundance Relative Abundance 70 65 60 55 50 70 65 60 8.70 8.16 45 35 8.59 35 50 40 40 30 8.37 8.18 8.70 8.35 8.6 8.84 25 8.8 8.2 8.4 T ime (m in) 8.6 8.8 8.0 8.5 T ime (min) lodta1 #37 RT : 8.48 AV: SB: 8.24-8.40 NL: 4.43E3 F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 368.9 100 9.0 T ime (min) lodta2 #37 RT : 8.48 AV: SB: 8.24-8.37 NL: 5.27E3 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 368.9 100 lodta4 #37 RT: 8.48 AV: SB: 27 7.67-8.18 , 8.84-9.03 NL: 5.03E3 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 368.9 100 90 80 80 80 70 70 70 60 50 40 30 206.9 Relative Abundance 90 Relative Abundance 90 Relative Abundance 8.16 30 20 8.4 8.67 35 20 8.2 8.59 40 8.56 25 8.86 25 7.88 55 45 45 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] M S lodta4 95 80 75 NL: 7.43E3 RT: 8.48 100 m /z= 368.5-369.5 F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] M S lodta2 95 80 Relative Abundance NL: 6.58E3 RT : 8.48 100 m /z= 368.5-369.5 F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS l odta1 95 30 RT : 7.79 - 9.43 RT : 8.04 - 8.82 RT : 8.08 - 8.89 60 50 40 206.9 30 294.0 206.9 60 50 40 30 294.0 20 350.0 10 20 350.0 10 207.6 293.4 220 240 260 280 300 m /z 320 340 360 293.9 350.0 10 20 220 240 260 280 300 m /z 320 340 360 207.7 220 240 260 280 300 m/z 320 340 360 60 40 20 80 60 40 20 293.9 8.0 8.2 8.4 8.6 8.8 9.0 250 60 40 20 80 60 40 20 294.0 8.0 8.2 8.4 8.6 8.8 9.0 250 100 349.9 60 80 60 40 20 80 60 40 20 293.9 100 8.2 8.4 8.6 8.8 9.0 250 80 60 40 20 0 18.5 350 60 40 294.0 8.2 8.4 8.6 T ime (min) 8.8 9.0 250 300 m /z 40 20 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g12 400 g12 #589 RT : 18.99 AV: NL: 2.68E4 F: + c SIM m s [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 80 356.1 60 40 20 19.0 19.5 350 T ime (m in) 80 8.0 NL: 3.72E4 349.9 350 100 RT : 19.00 19.002 80 60 NL: 4.72E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g14 40 20 18.5 g14 #589 RT : 18.99 AV: NL: 4.19E4 F: + c SIM m s [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 445.9 100 301.0 80 60 T ime (m in) 19.5 356.1 40 20 19.0 400 m /z RT : 18.50 - 19.50 20 356.0 60 m /z 20 Relative Abundance 100 g11 #589 RT : 18.99 AV: NL: 3.01E4 F: + c SIM m s [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 80 19.5 40 g14 #36 RT : 8.46 AV: SB: 20 8.05-8.30 , 8.75-9.00 NL: F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 RT : 8.49 8.485 RT : 19.00 19.004 60 m /z RT : 8.00 - 9.00 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g11 19.0 80 Relative Abundance T ime (min) 300 400 349.9 8.0 NL: 3.44E4 RT : 18.50 - 19.50 Relative Abundance 100 350 T ime (m in) g13 #36 RT : 8.46 AV: SB: 13 8.27-8.40 , 8.72-8.89 NL: F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 RT : 8.49 8.485 20 m /z 20 m /z RT : 8.00 - 9.00 40 19.5 40 18.5 350 RT : 19.00 19.003 80 Relative Abundance T ime (min) 300 356.0 60 19.0 RT : 18.50 - 19.50 Relative Abundance 80 g11 #589 RT : 18.99 AV: NL: 3.01E4 F: + c SIM m s [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 80 T ime (m in) g12 #36 RT : 8.46 AV: SB: 16 8.24-8.40 , 8.72-8.94 NL: F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 RT : 8.49 8.485 100 Relative Abundance 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g11 40 m /z RT : 8.00 - 9.00 Relative Abundance 300 60 Relative Abundance T ime (min) Relative Abundance 349.9 80 NL: 3.44E4 Relative Abundance 80 100 RT : 19.00 19.003 Relative Abundance Relative Abundance 100 RT : 18.50 - 19.50 Relative Abundance RT : 8.48 8.485 Relative Abundance RT : 8.00 - 9.00 Relative Abundance g11 #43 RT : 8.64 AV: SB: 20 8.05-8.30 , 8.75-9.00 NL: F: + c SIM m s [ 206.50-207.50, 293.50-294.50, 349.50-35 368.8 100 Relative Abundance M U GAN HEO + CHU N SALBUTAMOL ( 18.4 ng/ml) 301.7 350 400 m /z M U GAN HEO + CHU N SALBUTAMOL ( 9.2 ng/ml) 20 8.0 80 60 40 20 294.0 8.5 9.0 250 80 60 40 RT: 8.19 8.186 8.0 g22 #36 RT: 8.46 AV: SB: 17 8.13-8.32 , 8.70-8.92 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 80 60 40 293.9 9.0 250 Relative Abundance 100 80 60 NL: 1.22E5 TIC F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS g23 40 RT: 8.19 8.186 8.0 80 60 40 9.0 250 Relative Abundance 80 60 40 20 NL: 1.16E5 TIC F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ICIS g24 RT: 8.19 8.186 8.0 80 60 40 293.9 9.0 250 300 m/z 60 40 301.0 355.9 20 350 349.9 350 100 NL: 6.35E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g23 g23 #588 RT: 18.97 AV: SB: 28 18.73-18.94 , 19.17-19.39 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 80 60 301.0 356.0 40 20 19.0 RT: 19.00 18.998 80 60 19.5 350 400 m/z NL: 3.88E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g24 40 20 18.5 400 m/z RT: 18.50 - 19.50 20 80 19.5 20 g24 #36 RT: 8.46 AV: SB: 17 8.13-8.32 , 8.70-8.92 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 400 g22 #588 RT: 18.97 AV: SB: 28 18.72-18.93 , 19.17-19.38 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) 8.5 Time (min) m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g22 40 18.5 350 RT: 18.99 18.991 60 Relative Abundance RT: 8.49 8.486 350 19.0 80 m/z RT: 8.00 - 9.00 100 349.9 100 Relative Abundance Time (min) 300 20 Time (min) 8.5 NL: 3.35E4 RT: 18.50 - 19.50 293.9 356.0 40 m/z 20 g23 #36 RT: 8.46 AV: SB: 17 8.13-8.32 , 8.70-8.92 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 20 301.0 60 19.5 40 Relative Abundance RT: 8.51 60 18.5 350 RT: 19.00 19.001 80 m/z RT: 8.00 - 9.00 20 349.9 100 Relative Abundance Time (min) 300 80 19.0 RT: 18.50 - 19.50 20 g21 #588 RT: 18.98 AV: SB: 28 18.73-18.94 , 19.17-19.39 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) 8.5 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g21 40 Relative Abundance Relative Abundance NL: 7.46E4 TIC F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS g22 100 60 m/z RT: 8.00 - 9.00 RT: 8.49 300 80 Relative Abundance Time (min) 20 349.9 100 NL: 4.22E4 Relative Abundance 40 369.0 100 RT: 18.99 18.994 Relative Abundance 60 RT: 18.50 - 19.50 Relative Abundance 80 TIC F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ICIS g21 g21 #36 RT: 8.46 AV: SB: 17 8.13-8.32 , 8.70-8.92 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 g24 #589 RT: 18.98 AV: SB: 28 18.73-18.95 , 19.18-19.39 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Relative Abundance Relative Abundance 100 NL: 1.29E5 Relative Abundance RT: 8.49 8.485 Relative Abundance RT: 8.00 - 9.00 80 60 301.0 356.0 40 20 19.0 Time (min) 19.5 350 400 m/z M U GAN HEO + CHU N SALBUTAMOL ( 1.84 ng/ml) 20 60 40 293.9 8.5 9.0 250 80 60 40 20 g32 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 80 60 40 293.9 9.0 250 Relative Abundance 100 80 60 NL: 6.15E3 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS g33 40 20 80 60 40 294.0 349.9 250 Time (min) m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS g34 100 Relative Abundance NL: 5.85E3 80 60 300 40 20 350 80 60 40 293.9 349.9 8.5 Time (min) 9.0 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g32 250 300 m/z 80 356.0 60 40 20 19.5 350 RT: 19.00 19.001 60 NL: 5.24E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g33 40 20 100 350 g33 #588 RT: 18.97 AV: NL: 8.25E3 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 445.9 100 80 60 301.0 356.0 40 20 19.0 RT: 19.00 19.001 80 60 19.5 350 400 m/z NL: 7.08E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g34 40 20 18.5 400 m/z RT: 18.50 - 19.50 368.9 400 g32 #589 RT: 18.98 AV: NL: 3.39E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Time (min) 100 20 350 19.0 80 18.5 g34 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 Relative Abundance RT: 8.51 8.0 100 m/z RT: 8.00 - 9.00 NL: 4.43E4 RT: 18.50 - 19.50 9.0 20 Time (min) 8.5 40 m/z 20 g33 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.8 100 20 356.0 60 19.5 40 Relative Abundance RT: 8.49 60 18.5 350 RT: 19.00 19.000 80 m/z RT: 8.00 - 9.00 8.0 349.9 100 Relative Abundance Time (min) 300 80 19.0 RT: 18.50 - 19.50 20 g31 #589 RT: 18.98 AV: NL: 3.29E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Time (min) 8.5 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS g31 40 Relative Abundance Relative Abundance NL: 5.05E3 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS g32 100 60 m/z RT: 8.00 - 9.00 RT: 8.49 300 80 Relative Abundance Time (min) 8.0 349.9 Relative Abundance 8.0 80 20 100 NL: 3.62E4 Relative Abundance 40 368.8 RT: 19.00 19.001 Relative Abundance 60 100 RT: 18.50 - 19.50 Relative Abundance 80 g31 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 g34 #589 RT: 18.98 AV: NL: 5.21E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS g31 100 Relative Abundance NL: 4.78E3 Relative Abundance RT: 8.49 Relative Abundance RT: 8.00 - 9.00 80 60 356.0 40 20 19.0 Time (min) 19.5 350 400 m/z 40 20 8.0 8.27 80 60 40 20 293.9 8.5 9.0 250 80 60 40 20 80 60 40 293.9 250 80 60 NL: 3.11E4 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th13 40 20 60 40 9.0 250 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th14 100 80 60 40 20 8.19 80 60 40 294.0 349.9 300 m/z 350 8.5 Time (min) 9.0 250 80 301.0 60 356.1 40 20 350 NL: 2.48E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th13 th13 #589 RT: 18.98 AV: NL: 1.46E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 80 301.0 60 356.0 40 20 19.0 RT: 19.00 19.001 80 60 19.5 350 400 m/z NL: 3.28E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th14 40 20 18.5 400 m/z RT: 18.50 - 19.50 100 400 th12 #589 RT: 18.99 AV: NL: 2.52E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) th14 #41 RT: 8.59 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 20 350 19.5 20 Relative Abundance NL: 2.68E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th12 40 18.5 350 RT: 19.00 19.001 60 Relative Abundance RT: 8.49 300 20 19.0 80 m/z RT: 8.00 - 9.00 Relative Abundance 349.8 100 8.5 356.0 40 Time (min) 80 Time (min) 8.0 18.5 350 NL: 2.72E4 RT: 18.50 - 19.50 293.9 60 m/z 20 th13 #41 RT: 8.59 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 20 301.0 19.5 40 Relative Abundance Relative Abundance 100 60 m/z RT: 8.00 - 9.00 RT: 8.49 300 RT: 19.00 19.004 80 Relative Abundance Time (min) 8.0 349.9 100 9.0 80 19.0 RT: 18.50 - 19.50 20 th11 #589 RT: 18.99 AV: NL: 2.88E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) th12 #40 RT: 8.56 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 8.35 8.5 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th11 20 18.5 350 NL: 3.22E4 40 Relative Abundance Relative Abundance NL: 1.70E5 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th12 100 60 m/z RT: 8.00 - 9.00 RT: 8.48 300 80 Relative Abundance Time (min) 8.0 349.9 100 RT: 19.00 19.003 Relative Abundance 60 RT: 18.50 - 19.50 Relative Abundance 80 th11 #40 RT: 8.56 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th11 100 Relative Abundance NL: 1.98E5 th14 #590 RT: 19.00 AV: NL: 3.28E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Relative Abundance RT: 8.48 Relative Abundance RT: 8.00 - 9.00 Relative Abundance M U TH T HEO + CHU N SALBUTAMOL ( 18.4 ng/ml) 80 356.0 60 40 20 19.0 Time (min) 19.5 350 400 m/z 40 20 8.0 8.78 8.19 8.5 60 40 293.9 20 9.0 250 60 40 20 8.0 th22 #41 RT: 8.59 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 80 60 40 20 293.9 9.0 250 80 60 NL: 1.15E5 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th23 40 20 60 40 293.9 9.0 250 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th24 100 80 60 40 20 9.0 80 60 40 294.0 250 th22 #589 RT: 18.99 AV: NL: 4.20E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 445.9 100 301.0 80 60 356.0 40 20 350 400 m/z NL: 2.67E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th23 th23 #589 RT: 18.99 AV: NL: 2.67E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 80 60 301.0 356.1 40 20 19.0 19.5 350 400 m/z RT: 18.50 - 19.50 349.9 300 m/z 350 8.5 Time (min) 400 Time (min) th24 #42 RT: 8.62 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 20 350 19.5 20 Relative Abundance NL: 1.38E5 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th22 40 18.5 350 RT: 18.99 18.989 60 Relative Abundance RT: 8.51 300 20 19.0 80 m/z RT: 8.00 - 9.00 Relative Abundance 349.9 100 8.5 356.0 40 Time (min) 80 Time (min) 8.0 18.5 350 NL: 4.20E4 RT: 18.50 - 19.50 20 301.0 m/z 20 th23 #41 RT: 8.59 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 8.19 60 19.5 40 Relative Abundance Relative Abundance 100 60 m/z RT: 8.00 - 9.00 RT: 8.48 300 RT: 18.99 18.991 80 Relative Abundance Time (min) 8.0 349.9 100 8.5 80 19.0 RT: 18.50 - 19.50 8.19 th21 #588 RT: 18.98 AV: NL: 1.65E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) Relative Abundance Relative Abundance NL: 4.06E4 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th22 80 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th21 40 m/z RT: 8.00 - 9.00 100 300 60 Relative Abundance Time (min) RT: 8.49 349.9 80 NL: 5.42E4 Relative Abundance 60 80 100 RT: 18.99 18.992 Relative Abundance Relative Abundance 80 RT: 18.50 - 19.50 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th21 100 th21 #41 RT: 8.59 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 100 RT: 19.00 19.000 80 60 NL: 3.85E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th24 40 20 18.5 th24 #590 RT: 19.00 AV: NL: 3.85E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Relative Abundance NL: 3.90E4 RT: 8.51 Relative Abundance RT: 8.00 - 9.00 Relative Abundance M U TH T HEO + CHU N SALBUTAMOL ( 9.2 ng/ml) 80 60 356.0 40 20 356.7 19.0 Time (min) 19.5 350 400 m/z 40 8.68 20 8.19 8.0 60 40 20 293.9 8.5 9.0 250 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th32 Relative Abundance 100 80 60 40 8.19 8.79 th32 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 80 60 40 20 9.0 294.0 250 Relative Abundance 80 60 40 8.59 8.79 8.5 40 250 80 60 40 8.76 9.0 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th32 80 60 40 294.0 250 301.0 60 356.0 40 20 350 349.9 300 m/z 350 100 NL: 4.05E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th33 th33 #589 RT: 18.98 AV: NL: 3.30E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 80 356.1 60 40 20 19.0 RT: 19.00 19.001 80 60 19.5 350 400 m/z NL: 4.01E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th34 40 20 18.5 400 m/z RT: 18.50 - 19.50 20 80 19.5 20 th34 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 400 th32 #589 RT: 18.98 AV: NL: 4.61E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 445.9 100 Time (min) 8.5 Time (min) NL: 4.63E4 40 18.5 350 RT: 19.00 19.001 60 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th34 100 350 19.0 80 Relative Abundance NL: 5.65E3 RT: 8.49 Relative Abundance 349.9 100 m/z RT: 8.00 - 9.00 8.30 300 20 Time (min) 60 Time (min) 8.0 350 80 9.0 356.1 40 m/z RT: 18.50 - 19.50 294.0 60 20 th33 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 20 80 19.5 40 18.5 20 60 Relative Abundance NL: 8.35E3 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th33 100 8.21 300 RT: 19.00 19.000 80 m/z RT: 8.49 8.0 349.9 100 Relative Abundance Time (min) RT: 8.00 - 9.00 19.0 RT: 18.50 - 19.50 8.5 RT: 19.23 19.229 th31 #589 RT: 18.98 AV: NL: 6.07E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 445.9 100 301.0 Time (min) Relative Abundance NL: 1.06E4 RT: 8.49 20 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS th31 40 m/z RT: 8.00 - 9.00 8.0 300 60 Relative Abundance Time (min) 20 349.9 80 NL: 7.53E4 Relative Abundance 60 80 100 RT: 19.00 19.000 Relative Abundance Relative Abundance 80 RT: 18.50 - 19.50 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS th31 100 th31 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 th34 #589 RT: 18.98 AV: NL: 2.69E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Relative Abundance NL: 1.03E4 RT: 8.51 Relative Abundance RT: 8.00 - 9.00 Relative Abundance M U TH T HEO + CHU N SALBUTAMOL ( 1.84 ng/ml) 80 60 301.0 356.0 40 20 19.0 Time (min) 19.5 350 400 m/z 40 20 8.0 8.67 8.16 80 60 206.9 40 20 294.0 8.5 9.0 250 80 60 40 20 8.0 80 60 40 20 9.0 294.0 207.6 250 80 60 NL: 1.15E5 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta13 20 40 9.0 250 80 60 40 20 8.0 8.32 8.59 8.5 Time (min) 349.9 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta12 80 60 40 294.0 250 300 m/z ta12 #590 RT: 19.00 AV: NL: 2.20E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 80 356.1 60 40 20 350 NL: 2.06E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta13 349.9 350 80 301.0 60 356.1 40 20 19.0 RT: 18.99 18.991 80 60 19.5 350 400 m/z NL: 2.02E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta14 40 20 18.5 400 ta13 #589 RT: 18.99 AV: NL: 2.06E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) 100 400 m/z RT: 18.50 - 19.50 20 350 19.5 20 ta14 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 9.0 NL: 2.20E4 40 18.5 350 RT: 18.99 18.991 60 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta14 100 Relative Abundance NL: 1.08E5 20 19.0 80 m/z RT: 8.00 - 9.00 RT: 8.48 100 Relative Abundance Time (min) 300 356.0 40 Time (min) 60 293.9 60 m/z 20 18.5 350 80 301.0 RT: 18.50 - 19.50 20 80 19.5 40 ta13 #37 RT: 8.48 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 8.16 8.5 60 m/z 40 8.0 349.9 RT: 19.00 18.998 80 Relative Abundance Relative Abundance 100 300 100 Relative Abundance Time (min) RT: 8.48 19.0 RT: 18.50 - 19.50 RT: 8.00 - 9.00 RT: 19.20 19.201 ta11 #590 RT: 18.99 AV: NL: 3.41E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) ta12 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 369.0 100 8.16 8.5 20 Relative Abundance Relative Abundance NL: 1.21E5 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta12 100 40 RT: 18.76 18.758 m/z RT: 8.00 - 9.00 RT: 8.48 60 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta11 80 Relative Abundance Time (min) 300 100 349.9 NL: 3.54E4 RT: 19.00 19.003 Relative Abundance 60 RT: 18.50 - 19.50 Relative Abundance 80 ta11 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 369.0 100 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta11 100 Relative Abundance NL: 1.74E5 ta14 #589 RT: 18.99 AV: NL: 2.02E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Relative Abundance RT: 8.48 Relative Abundance RT: 8.00 - 9.00 Relative Abundance M U TH C ĂN GIA SÚC + CHU N SALBUTAMOL ( 18.4 ng/ml) 80 60 301.0 356.0 40 20 19.0 Time (min) 19.5 350 400 m/z 40 8.62 20 8.0 8.30 60 40 20 293.9 8.5 9.0 250 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta22 Relative Abundance 80 60 40 8.57 20 ta22 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 369.0 100 80 60 40 20 293.9 9.0 250 Relative Abundance 80 60 40 20 60 40 293.9 9.0 250 80 60 40 8.76 20 8.19 80 60 40 294.0 349.9 8.5 Time (min) 9.0 250 300 m/z 60 356.1 40 20 350 350 100 NL: 3.78E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta23 ta23 #589 RT: 18.99 AV: NL: 3.78E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 301.0 100 80 60 356.0 40 20 19.0 RT: 19.00 19.001 80 60 19.5 350 400 m/z NL: 3.07E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta24 40 20 18.5 400 m/z RT: 18.50 - 19.50 20 80 Time (min) ta24 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 400 ta22 #590 RT: 19.00 AV: NL: 2.40E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 19.5 20 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta24 100 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta22 40 18.5 350 RT: 18.99 18.993 60 Relative Abundance NL: 3.15E4 RT: 8.49 300 350 19.0 80 m/z RT: 8.00 - 9.00 Relative Abundance 349.9 100 8.5 20 Time (min) 80 Time (min) 8.0 18.5 350 NL: 2.40E4 RT: 18.50 - 19.50 20 356.1 40 m/z 20 ta23 #36 RT: 8.46 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 8.21 301.0 60 19.5 40 Relative Abundance NL: 5.84E4 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta23 100 60 m/z RT: 8.00 - 9.00 RT: 8.49 300 RT: 19.00 19.000 80 Relative Abundance Time (min) 8.0 349.9 100 8.5 80 19.0 RT: 18.50 - 19.50 8.24 ta21 #589 RT: 18.98 AV: NL: 1.98E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) Relative Abundance NL: 3.52E4 100 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta21 40 m/z RT: 8.00 - 9.00 RT: 8.49 300 60 Relative Abundance Time (min) 8.0 349.9 80 NL: 3.20E4 Relative Abundance 60 80 100 RT: 19.00 19.001 Relative Abundance Relative Abundance 80 RT: 18.50 - 19.50 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta21 100 ta21 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 ta24 #590 RT: 19.00 AV: NL: 3.07E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Relative Abundance NL: 3.18E4 RT: 8.49 Relative Abundance RT: 8.00 - 9.00 Relative Abundance M U TH C ĂN GIA SÚC + CHU N SALBUTAMOL ( 9.2 ng/ml) 80 60 356.1 40 20 19.0 Time (min) 19.5 350 400 m/z 40 20 8.0 8.70 8.35 60 40 20 294.0 8.5 9.0 250 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta32 Relative Abundance 80 60 40 20 8.68 8.19 ta32 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 80 60 40 293.9 9.0 250 80 60 NL: 4.13E3 m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta33 40 8.62 8.19 8.0 60 40 206.9 9.0 250 80 60 40 8.21 8.59 80 60 40 293.9 349.9 8.5 Time (min) 9.0 250 300 m/z 356.1 60 40 20 301.7 350 350 100 NL: 2.64E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta33 ta33 #590 RT: 19.00 AV: NL: 2.64E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 80 60 356.1 40 20 19.0 RT: 19.00 18.997 80 60 19.5 350 400 m/z NL: 2.56E4 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta34 40 20 18.5 400 m/z RT: 18.50 - 19.50 20 80 Time (min) ta34 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 400 ta32 #590 RT: 19.00 AV: NL: 3.61E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 301.0 100 19.5 20 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta34 100 Relative Abundance NL: 3.75E3 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta32 40 18.5 350 RT: 19.00 18.999 60 Relative Abundance RT: 8.49 300 350 19.0 80 m/z RT: 8.00 - 9.00 8.0 349.9 100 8.5 20 Time (min) 80 Time (min) 20 18.5 350 NL: 3.61E4 RT: 18.50 - 19.50 293.9 356.0 40 m/z 20 ta33 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 20 301.0 60 19.5 40 Relative Abundance Relative Abundance 100 60 m/z RT: 8.00 - 9.00 RT: 8.49 300 RT: 19.00 19.001 80 Relative Abundance Time (min) 20 350.0 100 8.5 80 19.0 RT: 18.50 - 19.50 20 ta31 #589 RT: 18.98 AV: NL: 3.14E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 Time (min) Relative Abundance NL: 6.00E3 100 20 18.5 350 m/z= 445.5-446.5 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] MS ICIS ta31 40 m/z RT: 8.00 - 9.00 RT: 8.49 300 60 Relative Abundance Time (min) 8.0 349.9 80 NL: 4.28E4 Relative Abundance 60 80 100 RT: 19.00 19.001 Relative Abundance Relative Abundance 80 RT: 18.50 - 19.50 Relative Abundance m/z= 368.5-369.5 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] MS ta31 100 ta31 #37 RT: 8.49 AV: SB: 21 8.10-8.32 , 8.59-8.89 NL: F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-35 368.9 100 ta34 #590 RT: 19.00 AV: NL: 2.56E4 F: + c SIM ms [ 300.50-301.50, 355.50-356.50, 445.50-446.50] 446.0 100 301.0 Relative Abundance NL: 6.20E3 RT: 8.49 Relative Abundance RT: 8.00 - 9.00 Relative Abundance M U TH C ĂN GIA SÚC + CHU N SALBUTAMOL ( 3.68 ng/ml) 80 60 356.1 40 20 19.0 Time (min) 19.5 350 400 m/z c:\xcalibur\data\beta agonist\6-11-07\m6 11/9/2007 6:27:15 PM M u M6_ Th c ăn cho heo phát hi n th y SAL 12ppb b ng GC/MS RT: 7.80 - 9.49 NL: 1.70E4 RT: 8.51 AA: 81118 SN: 271 100 m/z= 368.50-369.50 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] + m/z= 368.50-369.50 MS ICIS m6 90 70 60 50 40 30 368.94 100 90 80 Relative Abundance 80 Relative Abundance m6 #38 RT: 8.51 AV: SB: 22 8.10-8.38 , 8.65-8.92 NL: 8.24E3 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] 70 60 50 206.91 40 30 20 20 10 10 0 7.8 8.0 8.2 8.4 8.6 8.8 9.0 9.2 9.4 220 240 260 280 Time (min) NL: 1.84E4 RT: 8.54 AA: 137710 SN: 218 100 320 340 360 m6+std #39 RT: 8.54 AV: SB: 22 8.10-8.38 , 8.65-8.92 NL: 8.65E3 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] m/z= 368.50-369.50 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] + m/z= 368.50-369.50 MS ICIS m6+std 90 70 60 50 40 30 368.94 100 90 80 Relative Abundance 80 Relative Abundance 300 m/z RT: 7.80 - 9.49 206.90 70 60 50 40 30 20 20 10 10 7.8 294.03 349.97 8.0 8.2 8.4 8.6 8.8 9.0 9.2 9.4 220 240 260 280 Time (min) NL: 2.26E4 m/z= 368.50-369.50 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] + m/z= 368.50-369.50 MS s0,92c0,88 RT: 8.48 MA: 57391 SN: 49 100 300 320 340 s0,92c0,88 #36 RT: 8.45 AV: SB: 22 8.10-8.37 , 8.64-8.91 NL: 2.99E3 F: + c SIM ms [ 206.50-207.50, 293.50-294.50, 349.50-350.50, 368.50-369.50, 369.50-370.50] 80 70 60 50 368.83 100 90 80 Relative Abundance 90 70 60 50 40 206.93 30 294.01 20 349.97 10 40 8.2 8.3 8.4 8.5 Time (min) 360 m/z RT: 8.16 - 8.81 Relative Abundance 349.98 294.02 8.6 8.7 8.8 207.71 220 240 260 280 300 m/z 320 340 360 260 8.14 8.45 9.01 9.26 9.45 10.44 10.75 50 10 240 260 8.02 100 9.07 7.52 8.20 50 10.07 8.45 10.38 9.51 9 Time (min) 50 8.05 8.18 8.91 9.40 9.53 10.21 7.63 Time (min) 50 Time (min) 8.82 9.00 9.50 9.81 9.94 10.81 100 8.46 10 260 280 m/z 300 335.86 280 m/z 300 260 280 m/z 300 320 8.03 100 260 9.27 8.71 7.71 10.89 335.89 299.98 260 8.89 9.82 9.95 Time (min) 50 Time (min) 300 320 10 335.92 50 299.94 10.45 260 280 m/z 300 320 335.90 100 299.88 50 10 RT: 9.15 100 280 100 240 7.90 8.45 320 50 10 9.08 50 300 m/z 10.02 Time (min) 280 100 240 7.59 334.81 50 240 7.90 320 299.98 100 10.39 50 50 260 10 7.71 320 299.94 240 100 335.88 m/z 9.27 9.58 320 299.94 240 50 7.90 300 50 10 7.71 280 100 Time (min) 299.96 100 10 RT: 9.15 100 50 335.84 240 260 m/z 8.45 320 10 10.51 8.36 Relative Abundance 9.89 10.26 10.64 100 Relative Abundance 9.27 Relative Abundance Relative Abundance 8.83 300 100 50 240 8.01 m/z Relative Abundance Relative Abundance 8.34 280 299.92 Time (min) 260 50 10 7.46 335.79 240 50 100 320 50 10 7.71 7.90 300 299.99 100 Time (min) 100 m/z Relative Abundance Relative Abundance Time (min) 280 Relative Abundance 335.83 Relative Abundance 8 335.83 100 Time (min) 299.97 100 10.89 320 Relative Abundance 50 Relative Abundance Relative Abundance 7.71 300 m/z Time (min) 100 280 10.64 Relative Abundance 240 9.64 Relative Abundance 9.21 8.83 Relative Abundance 10.75 10 8.34 8.46 7.96 Relative Abundance 10.07 7.65 50 Relative Abundance 9.07 9.44 Relative Abundance 8.14 8.39 50 9.08 100 240 Relative Abundance 50 335.77 299.97 100 Relative Abundance 7.71 Relative Abundance 100 12/16/2008 4:44:07 AM 20 M u th c ph m ch bi n s n_CLEN Relative Abundance Relative Abundance e:\beta agonic\12-2008\15-12\1367a 260 280 m/z 300 320 334.81 299.98 100 50 240 260 280 m/z 300 320 240 260 8.27 7.89 10.81 9.07 8.58 9.32 9.88 Relative Abundance Relative Abundance 50 10.07 8.02 260 Relative Abundance Relative Abundance 10.08 10.39 100 50 Time (min) 9.08 10.07 10.57 9.89 10.57 260 10.82 Relative Abundance Relative Abundance Time (min) 300 300 100 7.89 10 280 m/z 10.07 10.81 100 260 9.71 9.95 100 10.76 9.08 9.39 9.95 10.38 300 320 260 Time (min) 280 m/z 300 320 335.82 100 50 300.03 240 260 280 300 320 335.80 100 299.92 50 240 260 280 300 320 m/z 50 335.88 300.01 10 RT: 9.15 100 320 m/z 7.71 8.14 8.39 300 50 10 50 280 100 240 50 7.90 8.21 8.83 9.15 335.84 50 10 7.72 320 m/z 8.27 Time (min) 300 299.97 240 9.51 334.81 260 10.76 10 9.07 280 100 Time (min) 50 240 10.51 8.57 50 320 299.98 100 260 Time (min) 50 280 9.08 299.91 260 240 320 335.84 240 m/z 9.64 10.13 m/z 50 280 100 10 RT: 9.15 8.89 300.02 Time (min) 100 8.33 50 320 335.91 50 300.02 10 7.96 m/z 9.08 9.20 300 100 240 Relative Abundance Relative Abundance 8.27 8.39 280 m/z 10.88 50 Time (min) 260 10.20 7.83 300.04 10 7.71 7.84 50 100 320 50 Time (min) 100 300 335.79 240 7.77 9.39 335.85 100 10 7.65 8.40 m/z Relative Abundance Relative Abundance 8.40 280 9.08 9.45 335.83 100 Time (min) 240 7.77 9.08 9.39 50 10 7.65 50 320 299.91 100 Time (min) 100 300 m/z Time (min) 100 280 8.21 8.45 Relative Abundance 10 50 Relative Abundance Relative Abundance 299.96 Relative Abundance 50 Relative Abundance 10.70 7.77 Relative Abundance 9.70 9.89 Relative Abundance 9.08 Relative Abundance 8.27 8.59 Relative Abundance 50 100 Relative Abundance 7.90 335.90 100 Relative Abundance 7.71 Relative Abundance 100 12/16/2008 1:05:55 AM 20 M u th c ph m ch bi n s n_CLEN Relative Abundance Relative Abundance e:\beta agonic\12-2008\15-12\1377a 10 334.81 299.98 100 50 240 260 280 m/z 300 320 250 8.42 9.04 10.60 9.35 9.60 50 293.95 10 250 8.98 10.41 Relative Abundance 10.10 9.54 Time (min) 8.80 10.36 9.36 293.93 9.36 7.74 8.05 9.54 10.10 10.60 8.80 10 293.96 250 Relative Abundance Relative Abundance 300 50 7.19 8.80 9.05 10 7.87 8.06 8.68 9.36 10.11 10.61 350.07 368.79 100 7.56 7.99 8.36 8.61 9.80 294.03 10.36 9.17 350.00 350 368.89 293.97 Relative Abundance 250 350 10.73 10.48 368.88 50 250 300 350 m/z 50 Time (min) 300 350.02 100 10 RT: 8.37 AA: 1211458 SN: 534 350 50 Time (min) 100 349.99 m/z 300 m/z 100 10 10.04 350 350 368.93 250 7.31 50 368.92 50 10 300 349.96 100 Time (min) 300 9.80 50 50 m/z 293.99 m/z 8.43 100 369.01 250 50 250 Time (min) 350 100 10 9.55 7.68 350 100 294.03 9.91 8.31 8.43 100 m/z 50 Time (min) 350.01 50 11 RT: 8.37 AA: 1211458 SN: 534 100 Time (min) 100 300 10.40 m/z 9.04 50 8.97 9.10 350 368.85 250 Relative Abundance Relative Abundance 8.36 349.95 368.84 10 10.16 8.66 50 300 350.05 368.95 m/z Time (min) 50 294.05 294.12 250 7.86 7.37 100 Time (min) 100 300 m/z 50 10 8.41 100 350 50 250 9.80 50 7.68 8.06 349.97 368.95 100 10 8.43 100 Relative Abundance 50 Relative Abundance Relative Abundance 8.36 7.92 m/z 8.23 7.37 300 10.67 100 Time (min) 100 Time (min) 100 Relative Abundance 7.80 7.37 9.74 8.93 350 Relative Abundance 50 Relative Abundance Relative Abundance 8.23 9.55 m/z Time (min) 100 300 8.43 Relative Abundance 10 7.87 7.19 Relative Abundance 350.02 368.83 Relative Abundance 293.86 50 Relative Abundance 50 Relative Abundance 8.67 8.79 Relative Abundance 10.41 9.60 8.24 100 Relative Abundance 10.10 8.42 7.92 Relative Abundance 7.80 100 Relative Abundance 8.23 100 50 12/16/2008 4:44:07 AM 20 m u th c ph m ch bi n s n_SAL Relative Abundance Relative Abundance e:\beta agonic\12-2008\15-12\1367a 10 369.00 100 50 293.99 349.99 250 300 m/z 350 9.60 7.37 10.60 10.41 50 10.16 9.04 7.86 293.96 250 10.11 10.60 9.05 Relative Abundance Relative Abundance 9.55 7.56 293.93 250 9.98 7.87 100 50 Time (min) 9.23 9.98 10.66 100 Time (min) 10 50 Time (min) 349.91 368.86 100 7.37 10 300 m/z 350.05 368.92 350.00 350 7.19 8.87 9.30 10.67 9.99 Time (min) 8.36 100 294.65 9.54 10.10 8.80 100 Time (min) 50 Time (min) 50 294.05 10.60 10 300 m/z 349.94 350 368.87 100 50 294.67 Relative Abundance 10 RT: 8.37 AA: 1211458 SN: 534 350 368.91 250 9.36 7.74 8.05 300 350.06 368.87 100 10 9.04 50 350 m/z 9.55 9.86 7.75 350.01 368.88 50 250 8.31 50 300 100 10 8.43 50 300 m/z 10.60 10.16 100 369.01 250 10.41 350 100 294.03 250 8.98 8.67 50 300 m/z 294.02 m/z 8.42 350 100 250 50 Time (min) 349.96 293.91 10.41 8.23 7.62 350 100 10 9.60 50 300 m/z 9.04 9.54 10.10 294.04 349.93 250 350 368.90 11 RT: 8.37 AA: 1211458 SN: 534 300 294.02 Time (min) 50 250 10.85 9.42 8.74 8.42 8.73 7.86 100 10 8.43 7.56 Relative Abundance 10.60 Relative Abundance Relative Abundance 9.55 Relative Abundance 9.36 10.11 Relative Abundance Relative Abundance 7.56 9.05 7.25 m/z 8.36 7.80 50 350 50 10 50 349.89 368.85 100 Time (min) 100 300 50 10 8.24 100 m/z 8.43 7.87 368.90 100 Time (min) 50 10 50 100 Time (min) 100 350 Relative Abundance 8.24 Relative Abundance Relative Abundance 8.36 100 300 m/z Time (min) 10.60 9.60 Relative Abundance 250 9.11 9.42 Relative Abundance 10 7.62 7.86 Relative Abundance 349.83 Relative Abundance 293.94 10.16 50 Relative Abundance 9.80 10.30 9.55 368.90 Relative Abundance 8.93 50 Relative Abundance 7.37 9.67 8.42 100 250 Relative Abundance 7.87 Relative Abundance 7.19 100 Relative Abundance 8.43 100 50 12/16/2008 1:05:55 AM 20 m u th c ph m ch bi n s n_SAL Relative Abundance Relative Abundance e:\beta agonic\12-2008\15-12\1377a 300 m/z 350.65 350 369.00 100 50 293.99 349.99 300 m/z 350 250 ... xuất phương pháp phân tích CLEN, SAL thịt heo, thức ăn nuôi heo, gan heo phương pháp sắc ký khí ghép khối phổ - Báo cáo đánh giá trạng tồn dư CLEN, SAL thịt heo, thức aên nuoâi heo, gan heo, . .. THU NGHIÊN C U PHƯƠNG PHÁP PHÂN TÍCH CÁC CH T KÍCH THÍCH TĂNG TRƯ NG H BETA-AGONIST (CLENBUTEROL, SALBUTAMOL) TRONG TH T HEO, GAN HEO, TH C ĂN NI HEO B NG PHƯƠNG PHÁP PHÂN TÍCH S C KÝ KHÍ GHÉP... dựng phương pháp phân tích Clenbuterol Salbutamol thịt heo, gan heo, thức ăn nuôi heo kỹ thuật sắc ký khí ghép khối phổ (GC/MS) Báo cáo hi n tr ng t n dư Clenbuterol Salbutamol th t heo, gan heo,