An ion chromatographic method was used to simultaneously determine nitrate and nitrite ions in biological samples. Ultrafiltration was used to produce a protein-free filtrate. Chloride interferences were eliminated by precipitation as the silver salt. Detection limits and average recoveries were 0.5 mg/L and 102% for nitrate and 0.2 mg/L and 78% for nitrite, respectively. Nitrate concentration was 2.1 +/- 1.8 mg/L and 4.9 +/- 0.8 mg/L in serum and ocular fluid of healthy cattle, respectively; nitrite was not detected. A severe case of nitrate poisoning in cattle was described and used to study the concentrations of nitrate and nitrite in samples obtained under natural conditions. Nitrate concentration of acutely poisoned cattle was 35% lower in ocular fluid at 158.1 +/- 51.4 mg/L, than in serum at 256.3 +/- 113.4 mg/L. Nitrite was not detected, because of the long processing time (> 3 hours) required for samples obtained in the field. A gradual decrease in ocular fluid nitrate of 29.4% at 24 hours, 25.9% at 36 hours, 51.6% at 48 hours, and 73.2% at 60 hours was observed; however, concentrations remained diagnostically significant (73.2 mg/L) 60 hours after death. Twenty-four hours after poisoning, the serum nitrate concentration of severely ill (52.7 +/- 51.9 mg/L) and moderately affected (12.4 +/- 5.7 mg/L) cattle that survived was indicative of the severity of clinical signs previously observed. Nitrate in serum and ocular fluid was stable in samples stored for 24 hours at 23 C, 1 week at 4 C, and 1 month at -20 C.

Tissue specimens from muscle, liver, kidney, and injection sites were collected, and serum was obtained from 3 calves euthanatized on each of posttreatment days 5 and 22. Calves were treated with 6.7, 13.4, or 20 mg of oxytetracycline (OTC)/kg of body weight, IM, once daily for 3 days; these dosages are 1, 2, and 3 times the label dose, respectively. One control calf was euthanatized on each of posttreatment days 5 and 22. In treated male calves killed 2 days after the last injection, OTC residues were detected in all tissues and serum, using high-performance liquid chromatography. Tissues from all injection sites also were considered positive for antimicrobial residues, using swab test on premises (STOP), microbial inhibition test (MIT), and thin-layer chromatography-biautography (TLCB) test. Kidney tissues from a calf given 13.4 mg of OTC/kg and kidney and liver tissues from a calf given 20 mg of OTC/kg also were considered positive, using the MIT and TLCB. Results of the STOP only were considered positive for the liver and kidney of a calf given 20 mg of OTC/kg, but substitution of Saskatoon antibiotic medium-3 for the original medium (antibiotic medium-5) allowed the STOP to detect residues in these tissues from all treated calves. In female calves killed 19 days after the last injection, the STOP, MIT, and TLCB procedures revealed positive results for tissues from some injection sites, but revealed negative results for other tissues. High-performance liquid chromatographic analyses detected OTC in tissues from injection sites from all treated calves, in muscle and liver from a calf given 20 mg of OTC/kg, and in kidneys from calves given 13.4 or 20 mg of OTC/kg. The STOP, MIT, and TLCB procedures lacked the sensitivity of high-performance liquid chromatography for detection of OTC residues.

This study was conducted to evaluate the ELISA kit for measuring the level of cortisol in the urine. The CV of within-run variation and day to day variation were 0.4-2.8 and 1.8-5.7, respectively. the minimum limitation of measurement was 1ng/ml. the cross reaction was high (CR50(%)=11.4-43.2) in prednisolone, 11-deoxycortisol, 21-deoxycortisol and predinosone. There was low and no cross reaction in other steroid. To develop the ELISA kit we measured the cortisol level in diluted urine with PBS (procedure I), extracted urine with methylene chloride (procedure II) and extracted methylene chloride-extracted urine from thin-layer chromatography (procedure III). The CV value of procedure I, II, III was 9.4-28.3%, 7.2-8.9% and 2.5-5.7%, respectively. There was significant difference between procedure I with II, and pro-cedure I with III(p0.01), but no difference between procedure II with III significantly(p0.01). The mean UCCR of urine collected through am 8 to 10 ws 9.5+_7.6(0.14-28.0) in 12-month-old dog(n

Biogenic amines (BAs) are formed as a result of specific free amino acid decarboxylation. Analysis of these metabolites may be of great importance to determine food quality and for monitoring the levels of biogenic amines such as histamine and tyramine related to intoxication episodes in humans. Chromatography is a chemistry separation technique used to characterize biogenic amines in foods. Variations of this technique (liquid, thin layer and gas chromatography) have been widely applied; however, the food matrix complex requires that changes in the methodology of extraction, derivatization and detection must be performed according to each group of foods. High-performance liquid chromatography is the most widely used chromatographic method applied for biogenic amines in foods. However, due to the current importance of biogenic amines in quality control and consumer safety, researchers try to develop new methods for a fast, reliable analysis of foods in the market. This review presents some chromatographic techniques applied to monitoring BAs in different foods of animal origin. | Aminas biogenicas sao formadas como resultado da descarboxilacao de aminoacidos livres especificos. A analise desses metabolitos e de grande importancia na determinacao da qualidade e monitoramento de biogenicas como histamina e tiramina relacionadas com episodios de intoxicacao em humanos. A cromatografia e uma tecnica de separacao química usada para caracterizar aminas biogenicas. Variacoes da tecnica (cromatografia liquida, em camada delgada e gasosa) tem sido amplamente usadas, porem a complexidade da matriz alimentar faz com que sejam realizadas mudancas nos processos de extracao, derivatizacao e deteccao em concordancia com cada grupo de alimento. A cromatografia liquida de alta eficiencia (CLAE) e o metodo mais utilizado na determinacao de aminas biogenicas em alimentos. Contudo, devido a importancia das aminas biogenicas no controle da qualidade e a seguranca do consumidor, os pesquisadores tentam desenvolver novos metodos com o intuito de uma analise mais rapida e precisa para o controle de alimentos no mercado. O objetivo da revisao e apresentar algumas tecnicas cromatograficas aplicadas no monitoramento de aminas biogenicas em produtos de origem animal.

Objective of the present study was to purify hCG contained in commercial preparation (Pregnyl® - Organon) by affinity chromatography and to evaluate ovulation rates in cows injected with the purified hormone. A chromatography column containing concanavalin-A sepharose (Con-A) matrix was equilibrated and one hundred thousand international units (UI) of Pregnyl® were applied to the column. Fractions of 3,8µL were collected (4°C) every 5 minutes for 12,5h, to yield a total of 150 fractions. After fraction 76, a column buffer containing 0,3M ±-methylglicoside was added to the column. Protein concentrations were estimated by espectrophotometry (280nm). Two peaks of absorbance were observed, between fractions 14 and 19, and fractions 90 and 100. Fractions 14-19 and 90-150 were sequentially grouped in pairs concentrated by ultrafiltration and analyzed by SDS-PAGE. The first peak of absorbance resulted from proteins contained in the commercial product, eluted from the column and not adsorbed by the Con-A matrix. The second peak represented proteins adsorbed by the column, especially hCG. The fractions containing hCG were grouped in a single sample and protein concentration was measured by the Lowry method. Cows in day 5 of the estral cycle, which had a dominant follicle larger than 8mm, were injected with 0,1, 0,2 or 0,3mg protein of the purificated sample. The ovulation rate 48 hours after treatment was 0%, 50% and 75%, respectively. It was demonstrated that relative concentration of hCG can be increased by affinity chromatography and biological activity of purified hCG was satisfactory preserved. | O objetivo do presente estudo foi purificar o hCG contido em uma preparação comercial (Pregnyl® Organon) por cromatografia de afinidade e avaliar a taxa de ovulação de vacas injetadas com o hormônio purificado. Uma coluna de cromatografia contendo matriz de Concanavalina-A Sepharose (Con-A) foi equilibrada e cem mil unidades internacionais (UI) de Pregnyl® foram aplicadas na coluna. Frações de 3,8µL foram colhidas (4ºC) a cada 5 minutos durante 12,5 horas, resultando em um total de 150 frações. Após a fração 76 adicionou-se à coluna solução tampão contendo 0,3M de ±-metilglicosidase. A concentração protéica das frações foi estimada por espectrofotometria (280nm). Foram observados dois picos de absorbância, um entre as frações 14 e 19 e outro entre as frações 90 e 100. As frações 14 a 19 e 90 a 150 foram reunidas seqüencialmente aos pares, concentradas por ultrafiltração e analisadas por SDS-PAGE. O primeiro pico resultou da eluição das proteínas não adsorvidas pela matriz de Con-A, enquanto o segundo representou as proteínas adsorvidas pela coluna, especialmente o hCG. As frações contendo hCG foram reunidas em uma única amostra cuja concentração protéica foi quantificada pelo método de Lowry. Vacas no 5º dia de um ciclo estral sincronizado, apresentando um folículo dominante maior que 8mm de diâmetro, foram injetadas com 0,1, 0,2 ou 0,3mg de proteína contida na amostra purificada. Após 48 horas, a taxa de ovulação observada foi de 0%, 50% e 75%, respectivamente. Foi demonstrado no presente estudo que a técnica de cromatografia por afinidade foi eficiente para aumentar a concentração relativa do hCG preservando sua atividade biológica.

O objetivo do presente estudo foi purificar o hCG contido em uma preparação comercial (Pregnyl® Organon) por cromatografia de afinidade e avaliar a taxa de ovulação de vacas injetadas com o hormônio purificado. Uma coluna de cromatografia contendo matriz de Concanavalina-A Sepharose (Con-A) foi equilibrada e cem mil unidades internacionais (UI) de Pregnyl® foram aplicadas na coluna. Frações de 3,8µL foram colhidas (4ºC) a cada 5 minutos durante 12,5 horas, resultando em um total de 150 frações. Após a fração 76 adicionou-se à coluna solução tampão contendo 0,3M de ±-metilglicosidase. A concentração protéica das frações foi estimada por espectrofotometria (280nm). Foram observados dois picos de absorbância, um entre as frações 14 e 19 e outro entre as frações 90 e 100. As frações 14 a 19 e 90 a 150 foram reunidas seqüencialmente aos pares, concentradas por ultrafiltração e analisadas por SDS-PAGE. O primeiro pico resultou da eluição das proteínas não adsorvidas pela matriz de Con-A, enquanto o segundo representou as proteínas adsorvidas pela coluna, especialmente o hCG. As frações contendo hCG foram reunidas em uma única amostra cuja concentração protéica foi quantificada pelo método de Lowry. Vacas no 5º dia de um ciclo estral sincronizado, apresentando um folículo dominante maior que 8mm de diâmetro, foram injetadas com 0,1, 0,2 ou 0,3mg de proteína contida na amostra purificada. Após 48 horas, a taxa de ovulação observada foi de 0%, 50% e 75%, respectivamente. Foi demonstrado no presente estudo que a técnica de cromatografia por afinidade foi eficiente para aumentar a concentração relativa do hCG preservando sua atividade biológica.