Oxytetracycline analysis

The overall biosynthetic pathway to the tetracychnes has been reviewed (74). Studies (75—78) utilising labeled acetate and malonate and nmr analysis of the isolated oxytetracycline (2), have demonstrated the exclusive malonate origin of the tetracycline carbon skeleton, the carboxamide substituent, and the folding mode of the polyketide chain. Feeding experiments using [1- 02] acetate and analysis of the nmr isotope shift effects, led to the location of... 

United States Pharmacopoeia 28 [1] describes a microbiological method under antibiotics-microbial assays for the analysis of OTC and nystatin capsules, OTC and nystatin for oral suspension, OTC HC1 and hydrocortisone ointment, and OTC HC1 and polymyxin B sulfate ointment. The methods are relative rather than absolute, which are based on the determination of the level of oxytetracycline by a microbiological response to a series of standard oxytetracycline concentrations by a... 

HPLC methods have been widely used for the analysis of OTC in different samples. As described above in the Section 2.3, the HPLC method is described in most of compendia [1,2,4,7] for determination of OTC in bulk drug substances and in some pharmaceutical preparations. The application of HPLC methods for the analysis of antibiotics including oxytetracycline has been recently reviewed by Diaz-Cruz et al. [37] and Lunn [38], A summary of HPLC method for the analysis of OTC is presented in Table 3. 

Table 2. Analysis of oxytetracycline by TLC (data cited from CBS 1.09, 2003, Camag)...

Table 3. Summary of HPLC conditions for the analysis of oxytetracycline... 

For confirmatory assay, liquid chromatography-tandem mass spectrometry (LC-MS/MS) is becoming more frequently used in the analysis of OTC owing to its high sensitivity and ability. Electrospray ionization (ESI) [55-57] and atmospheric pressure chemical ionization (APCI) [41] methods combined with tandem mass spectrometry are favored because of their higher sensitivity and better reproducibility. Hamscher et al. [58] developed a method for the determination of persistent TC residues in soil fertilized with manure by HPLC tandem mass spectrometry, MS-MS, and confirmation by MS-MS-MS. Zhu et al. [59] developed an LC-tandem mass spectrometry for the analysis of common tetracyclines in water. The detection limit for oxytetracycline was 0.21 pg/L. Lykkeberg et al. [60] used LC-MS/MS for determination of oxytetracycline and its impurities EOTC, TC, ETC, ADOTC, oc-AOTC, and /i-AOTC. 

An important area of direct biochemical interference is that caused by fluorescent or fluorescent quenching materials in the blood or urine after the administration of a drug. These interferences may be observed during catecholamine analysis in urine from patients receiving -methyldopa, tetracyclines, chlortetracyclines, oxytetracycline, erythromycin, chlorpro-mazine, or quinidine (A2, G5). 

Tiamulin is well absorbed when given orally, and is excreted mainly in bile within 28 h. The withdrawal period after oral administration to pigs is 5 days, to allow for excretion of metabolites. However, when tiamulin is administered in combination with oxytetracycline, the elimination of tiamulin residues from tissues is slightly prolonged, thus maintenance of a withdrawal period of 2 weeks is required from the public health point of view, based on an analysis of the elimination profile (259). 

Analysis of the 1994 tissue residue data revealed that FSIS reported 2514 animals containing violative residues. FDA, in cooperation with participating states, conducted follow-up investigation on 1076 (45%) of the reported violations. The drugs most frequently identified as causing antibiotic residues included penicillin (21%), oxytetracycline (10%), sulfamethazine (10%), streptomycin (6%), tetracycline (5.2%), neomycin (4.1%), gentamicin (3.7%), and sulfadimeth-... 

With the differential pulse polarography [245], the antibiotics can be determined at low concentration, if necessary, at the ppm or even sub-ppm level. Tetracycline hydrochloride is determined in aqueous acetate buffer pH 4 (detection limit 0.1 ppm), but for the analysis of chlortetracycline hydrochloride, oxytetracycline hydrochloride and free tetracycline, a non-aqueous medium must be used. Streptomycin sulphate is analysed in alkaline solution, trace quantities of zinc being masked by Na2EDTA, and the detection limit is 1 ppm. A determination in blood serum or urine is also possible but the peak potentials are shifted here to more negative values. The polarographic determination is preceded by ultrafiltration. Penicillin G potassium and ampicillin must be first functionalised by nitrosation. The authors also recommend an analysis of mixtures which is however demonstrated only with chloramphenicol and tetracycline, at 2.4 and 4.2 ppm, respectively. 

Figure 14.8 LC-ESI-MS chromatogram in time-scheduled SIM mode for the analysis of milk spiked with 25 pg/kg of tetracyclines. Peaks MC=minocycline, TC=tetracycline, OTC=oxytetracycline, CTC=chlortetracycline, and DC=doxy-cycline. Reprinted from [15] with permission. 2002, John Wiley and Sons Ltd.

Fig. 18 a-c. Purification and ESMS analysis of oxytetracycline acyl carrier proteins after growth and induction in minimal medium containing 5NH4C1 a Q-Sepharose FPLC trace of E. coli protein fractions containing ACP b ESMS of N-oxytetracycline apo-ACE note distinctive envelope of Na+ adducts c ESMS of N-oxytetracycline ho/o-ACP, again showing Na+ adducts, as well as a significant proportion (ca. 10%) of unlabelled holo-ACP... 

Croubles, S. Baeyens, W. Van Peteghem, C. Evaluation of a narrow-bore HPLC column for trace level analysis of tetracyclines-a comparison with a conventional column. Biomed.Chromatogr., 1995, 9, 251-253 [simultaneous chlortetracycline, demeclocycline, oxytetracycline]... 

Craigmill AL, Miller GR, Gehring R, Pierce AN, Riviere JE, Meta-analysis of pharmacokinetic data of veterinary drugs using the Food Animal Residue Avoidance Databank Oxytetracycline and procaine peniciUin G, J. Vet. Pharmacol. Ther. 2004 27 343-353. 

Stolker et al. " described an analytical method based on TFC-LC-MS/MS for the direct analysis of 11 veterinary drugs (belonging to seven different classes) in milk. The method was applied to a series of raw milk samples, and the analysis was carried out for albendazole, difloxacin, tetracycline, oxytetracycline, phenylbutazone, salinomycin-Na, spiramycin, and sulfamethazine in milk samples with various fat contents. Even without internal standards, results proved to be linear and quantitative in the concentration range of 50-500 (xg/1, as well as repeatable (RSD<14% sulfamethazine and difloxacin <20%). The limits of detection were between 0.1 and 5.2 xg/l, far below the maximum residue limits for milk set by the EU. While matrix effects, namely, ion suppression or enhancement, were observed for all the analytes, the method proved to be useful for screening purposes because of its detection limits, linearity, and repeatability. A set of blank and fortified raw milk samples was analyzed and no false-positive or falsenegative results were obtained. 

Goto et al. " described a simple, rapid, and simultaneous analysis method for oxytetracycline, tetracycline, chlortetracycline, penicillin G, ampicillin, and nafcillin in meat using electrospray ionization tandem mass spectrometry. The samples were homogenized with water followed by a centrifugal ultrafiltration after addition of internal standards (demeclocycline, penicillin G-ds, ampicillin-fi 5, and... 

Cherlet M, De Baere S, De Backer P, Quantitative analysis of oxytetracycline and its 4-epimer in calf tissues by liquid chromatography combined with positive electrospray ionization mass spectrometry, Ana/yi f 2003 128 871-878. 

AO AC, Chlortetracycline, oxytetracycline and tetracycline in edible animal tissues, 995.09, in Official Methods of Analysis, 18th ed. (revised), AO AC International, Gaithersburg, MD (available at http //www.eoma.aoac.org/gate way/readFile. asp id=995 09. pdf accessed 3/24/10). 

Stehly GR, Gingerich WH, Kiessling CR, Cutting JH, A bridging study for oxytetracycline in the edible fillet of rainbow trout Analysis by a liquid chromatographic method and the official microbial inhibition assay, J. AO AC Int. 1999 82(4) 866-870. 

Kim ES, Bibb MJ, Butler MJ, Hopwood DA, Sherman DH. Sequences of the oxytetracycline polyketide synthase encoding otc genes from Streptomyces nmosio. Gene 1994 141 141-142. Khosia C, McDaniel R, Ebert-Khosla S, Sherman DH, Bibb MJ, Hopwood DA. Genetic construction and functional analysis of hybrid polyketide synthases containing heterologous acyl carrier proteins. J Bacteriol 1993 175 2197-2204-... 

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