Background. The European Union prohibits the use of chloramphenicol (CAP) as a veterinary drug in food-producing animals. Nevertheless, CAP have been detected in milk products (liquid milk and milk powder). Therefore, it is necessary to develop sensitive methods for determining CAP residues in milk powder. Objective. The aim of this study was to develop and validate a confirmatory method for determination of CAP in milk powder. Material and methods. Chloramphenicol was determined in milk powder using LC-ESI-MS/MS in negative mode. After fat removing milk powder sample was extracted/cleaned-up with a Chem Elut extraction cartridge. Separation was achieved on a Phenomenex Luna C-18 column with acetonitrile-water as a mobile phase. The mass spectrometer was operated in multiple reaction monitoring mode (MRM). Four transitions were monitored m/z 321→152, 321→194, 321→257 (CAP) and 326→157 (IS CAP-d5). Results. Linearity, accuracy, precision, decision limit (CCa), detection capability (CCb) and ruggedness were determined for m/z 321→152. The mean relative recoveries (inter standard-corrected) of CAP from whole milk powder spiked at levels 0.1, 0.2, 0.3 and 0.6 mg/kg were in the range 95 - 103%. Relative standard deviation (RSD%) of recoveries at all spiked levels were less than 14%. RSDs within-laboratory reproducibility calculated at fortification of 0.3 mg/kg was less than 16%. CCa and CCb were below 0.1 mg/kg. Conclusions. The developed LC-MS/MS method allows the determination of CAP in milk powder. The method was validated according to the Commission Decision No. 2002/657/EC requirements. This method can be applied to determination CAP in whole and skim milk powder.
Analytical methods based on gas chromatography- mass spectrometry (GC-MS/MS) and liquid chromatography- mass spectrometry (LC-MS/MS) were developed for the determination of chloramphenicol (CAP) in milk. GC-MS/MS was performed in negative chemical ionization (NCI) mode by monitoring the transitions - m/z 466→304, 466→322, and the results were compared with LC-MS/MS, in electrospray mode by monitoring the transitions of all the selected ions m/z 321→152, 321→257. The decision limit (CCα) for CAP determination by LC-MS/MS was established at a level of 0.11 µg/kg, while the corresponding value for GC-MS/MS was 0.083 µg/kg. Detection capability (CCß) for CAP by LC- MS/MS was 0.15µg/kg and for GC-MS/MS was 0.14 µg/kg. As it was found, both methods are useful for CAP determination in milk.
The method was developed for the determination of chloramphenicol (CAP) in animal urine using LC-ESI-MS- MS. The sample was diluted with water, loaded onto a Chem Elut cartridge, and eluted with ethyl acetate. LC separation was achieved by using a Phenomenex Luna C-18 column. The mass spectrometer was operated in multiple reaction monitoring mode (MRM) with negative electrospray interface (ESI-). The four transitions were monitored m/z 321—>257, 321—> 194, 321—> 152, 326—> 157(IS) and for quantification, the transition m/z 326—>152 was chosen. The method validation was done according to the criteria laid down in Commission Decision No. 2002/657/EC. The validation includes the determination of specification, linearity, repeatability, within-laboratory reproducibility, accuracy, trueness, decision limit (CCα), and detection capability (CCβ). Urine_samples were fortified at CAP levels 0.30, 0.45, and 0.60 µg/kg with CAP-d5 as internal standard. At these levels, repeatability (RSD %) was lower than 9%, accuracy (RE, %) ranged from -8.7% to 0.4% and mean trueness (recovers) was between 91% and 100%. RSD within-laboratory reproducibility was lower than 10%. The limit of decision (CCα) and detection capability (CCβ) was 0.08 and 0.10 µg/kg.
W pracy przedstawiono metodę potwierdzającą oznaczania chloramfenikolu w tkance mięśniowej zwierząt przy zastosowaniu techniki LC-MS/MS spełniającej zalecenia Decyzji Komisji nr 2003/181/WE i nr 2002/657/WE.
Przedstawiono metodę oznaczania chloramfenikolu w mleku proszku. Próbki były oczyszczane przy zastosowaniu polimerów z odwzorowaniem cząsteczkowym (МІР). Analizę przeprowadzono w układzie LC-ESI-MS/MS z zastosowaniem kolumny Luna C 18 Phenomenex. Jako standard wewnętrzny zastosowano CAP-d5. Metodę zwalidowano zgodnie z kryteriami decyzji Komisji nr 2002/657/WE. Średni odzysk próbek wzmocnionych na poziomie 0,3 µg/kg mieścił się w zakresie 104-111%. Limit decyzyjny (CCα) i zdolność wykrywania (CCß) wynosiły odpowiednio 0,06 µg/kg i 0,09 µg/kg.
In Penicilium isariiforme there occur two endogenous rhythms: the growth-“wave” rhythm and sporulation rhythm in the form of coremia-bearing zonations. The growth rhythm of wave type occurring in P. isariiforme in light and in darkness is an endogenous spontaneous rhythm, whereas the sporulation rhythm manifested by the formation of spore-bearing coremia zonations represents an analogue induced rhythm. For this rhythm to occur a light impulse is necessary. The growth-wave rhythm has a long period 120 h, the sporulation rhythm has a 30 h period. Asparagine seems to regulate the sporulation rhythm; but it does not exert any greater influence on the wave rhythm in cultures grown in light nor in darkness. An addition of asparagine shortened the period of first zonation by a few hours, and the successive sporulation periods followed within very short time. Inhibitors such as: avidin. chloramphenicol, cycloheximide and puromycine controlled the length of the period of the sporulation rhythm.
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