Quantificação de fatores de crescimento na pele de equinos tratada com plasma rico em plaquetas

O plasma rico em plaquetas (PRP) é um produto derivado da centrifugação do sangue total, sendo rico em fatores bioativos, como os de crescimento. Apesar da ampla utilização em processos cicatriciais, há controvérsia sobre a eficácia da terapia na cicatrização cutânea. O objetivo desse estudo foi quantificar e comparar a concentração dos fatores TGF-β1 e PDGF-BB no PRP, plasma sanguíneo e pele, durante diferentes fases do processo de cicatrização da pele tratada ou não com PRP. Foram utilizados sete equinos machos castrados, mestiços, hígidos, com idade entre 16 e 17 (16,14±0,63) anos. Três lesões em formato quadrangular (6,25cm²) foram produzidas cirurgicamente nas regiões glúteas direita e esquerda de todos os animais. Doze horas após indução das feridas, 0,5mL do PRP foi administrado em cada uma das quatro extremidades das feridas de uma das regiões glúteas (Grupo tratado = GT), escolhida aleatoriamente. A região contralateral foi utilizada como controle (GC). As feridas foram submetidas à limpeza diária com água Milli Q, e amostras foram obtidas mediante biópsias realizadas com Punch de 6mm. Foram obtidas seis biópsias de pele, sendo a primeira realizada logo após a produção da ferida (T0), e as demais com 1 (T1) 2 (T2) 7 (T3) e 14 (T4) dias após a indução da lesão. A sexta biópsia (T5) foi obtida após completo fechamento da pele, que ocorreu aproximadamente aos 37 dias (36,85±7,45, GC; 38,85±6,46, GT). Também foram obtidas amostras de sangue com EDTA em todos os tempos mencionados. A quantificação dos fatores de crescimento TGF-β1 e PDGF-BB na pele, PRP e plasma sanguíneo foi realizada pela técnica ELISA. Os dados foram analisados estatisticamente pelo teste t, correlação de Pearson e regressão, utilizando nível de significância de 5%. Não houve diferença entre os grupos, nos valores dos dois fatores de crescimento mensurados na pele, nos diferentes tempos. Também não houve correlação entre a quantidade dos fatores de crescimento presentes na pele e no plasma. Por outro lado, correlação positiva foi observada entre PRP e pele no grupo tratado, para os fatores de crescimento TGF-β1 (r=0,31) e PDGF-BB (r=0,38), bem como entre ambos os fatores de crescimento presentes no PRP (r=0,81). Considerando as concentrações dos fatores de crescimento no T0, os maiores valores cutâneos (p<0,05) do TGF-β1, em ambos os grupos, ocorreram nos tempos T3 e T5. Valores mais elevados (p<0,05) do PDGF-BB ocorreram no T4 (GT) e T5 (GC). No plasma não houve alteração nas concentrações desses fatores em relação ao T0, o que sugere que o PRP não acarreta efeito sistêmico, quando os procedimentos adotados na presente pesquisa são utilizados. A administração local de PRP no volume estudado, 12 h após indução cirúrgica de ferida cutânea na região glútea de equinos não ocasiona maiores concentrações dos fatores de crescimento TGF-β1 e PDGF-BB no plasma sanguíneo e pele, durante o processo de cicatrização.

Babesia bovis: expression of adhesion molecules in bovine umbilical endothelial cells stimulated with plasma from infected cattle

Ten male, 12-month-old Jersey with intact spleens, serologically and parasitologically free from Babesia were housed individually in an arthropod-free isolation system from birth and throughout entire experiment. The animals were randomly divided into two groups. Five animals (group A) were intravenously inoculated with 6.6 X10(7) red blood cells parasitized with pathogenic sample of Babesia bovis (passage 7 BboUFV-1), for the subsequent "ex vivo" determination of the expression of adhesion molecules. Five non-inoculated animals (group B) were used as the negative control. The expression of the adhesion molecules ICAM-1, VCAM, PECAM-1 E-selectin and thrombospondin (TSP) was measured in bovine umbilical vein endothelial cells (BUVECs). The endothelial cells stimulated with a pool of plasma from animals infected with the BboUFV-1 7th passage sample had a much more intense immunostaining of ICAM-1, VCAM, PECAM-1 E-selectin and TSP, compared to the cells which did not received the stimulus. The results suggest that proinflammatory cytokines released in the acute phase of babesiosis may be involved in the expression of adhesion molecules thereby implicating them in the pathophysiology of babesiosis caused by B. bovis.

Padronização de técnica manual para obtenção de plasma rico em plaquetas de bovino

Resumo: Para padronização de uma técnica manual para a obtenção de plasma rico em plaquetas (PRP) autólogo em bovinos com custo reduzido (método manual) e de boa qualidade (capacidade de concentrar plaquetas, alta concentração de fatores de crescimento e contaminação reduzida com leucócitos e eritrócitos), que poderá ser utilizado como um agente modulador da resposta imune de vacas com diferentes enfermidades, 450 ml de sangue total de nove vacas clinicamente saudáveis e com perfil hematológico normal foi coletado em bolsas de sangue CPDA-1 e processado dentro de quatro horas após a coleta. O sangue foi separado em alíquotas para avaliar 8 protocolos (P) de centrifugação dupla que variaram quanto a velocidade e o tempo de centrifugação. A contagem de plaquetas, eritrócitos e leucócitos na suspensão obtida (PRP) foi realizada pelo método manual em câmara de Neubauer: P5 (400g e 800g ambos durante 10 min) foi o protocolo com maior número de plaquetas, seguido por P3 (120g e 473g ambos durante 10 min), P4 (300g e 640g durante 10 min cada), P6 (640g durante 10 min e 640g durante 5 min), P8 (640g durante 5 min e 120g durante 10 min) e P7 (720g e 720g durante 5 min) e diferentes (p<0,05) dos menores valores encontrados em P1 (120g e 240g, ambos por 5 minutos) e P2 (120g e 473g ambos por 5 min). Em relação aos eritrócitos, P8, P7, P6, P5 e P4 apresentaram menores concentrações e maiores valores (p<0,05) foram observados em P3 e P2. Menores quantidades de leucócitos foram observadas em P5, P6, P8 e P7 com o maior valor obtido em P2 (p<0,05). Todos os protocolos (P1 a P8) foram eficientes em concentrar plaquetas sendo o valor mais baixo (3,65±0,79) observado em P1. Em relação aos fatores de crescimento ao se mensurar TGF- 1, os protocolos P1 e P8 evidenciaram valores mais elevados. De acordo com os resultados obtidos os protocolos P5 e P8 apresentaram os melhores resultados para confecção de PRP bovino.

Clinical, hematological, total plasma protein and fibrinogen parameters of magellanic penguins ( Spheniscus magellanicus ) pre- and post-rehabilitation

Abstract: Magellanic penguins (Spheniscus magellanicus) usually arrive in poor body conditions at Brazilian beaches during the winter. Hematology provides valuable information about clinical and immunity status of the animals. The aims of this study were to determine the hematologic, total plasma protein (TPP) and fibrinogen profiles of young and adult magellanic penguins in PROAMAR and CETAS-SC, relating these results with the state of health and survival possibility of the animals. In Paraná 14 animals were evaluated in pre and eight in post-rehabilitation and 29 animals were evaluated in Santa Catarina after rehabilitation. Before rehabilitation, all animals showed weakness. In hematological exams of these animals, we found that anemia was present in 83% of the penguins that died and 50% of those which survived. The heterophils/lymphocytes (H/L) ratio was 3.87±0.57 in animals that died, significantly higher than the average of 2.20±0.30 for animals that survived. These two parameters are useful to assess the survival possibility of animals to rehabilitation. The body condition score was positively correlated with hematocrit and TPP, and negatively correlated with H/L ratio. After rehabilitation, the values were similar to other animals of the family Spheniscidae, with averages ranging from 1.64 to 1.90x106 erythrocytes/μL; 43.38 to 48.80% of hematocrit; 12.45 to 13.52g/dL of hemoglobin; 8,684 to 14,011 leukocytes/μL; 4,767 to 8,041 heterophils/μL; 3,215 to 4,951 lymphocytes/μL; 95 to 655 eosinophils/μl; 179.8 to 277.9 monocytes/μL; 141 to 184.9 basophils/μL; and 1.26 to 1.74 of H/L ratio. These parameters can therefore be used as reference values and release parameters for young and adult Magellanic penguins in captivity on the rehabilitation centers.

Method to obtain platelet-rich plasma from rabbits (Oryctolagus cuniculus )

Abstract: Platelet-rich plasma (PRP) is a product easy and inxpesnsive, and stands out to for its growth factors in tissue repair. To obtain PRP, centrifugation of whole blood is made with specific time and gravitational forces. Thus, the present work aimed to study a method of double centrifugation to obtain PRP in order to evaluate the effective increase of platelet concentration in the final product, the preparation of PRP gel, and to optimize preparation time of the final sample. Fifteen female White New Zealand rabbits underwent blood sampling for the preparation of PRP. Samples were separated in two sterile tubes containing sodium citrate. Tubes were submitted to the double centrifugation protocol, with lid closed and 1600 revolutions per minute (rpm) for 10 minutes, resulting in the separation of red blood cells, plasma with platelets and leucocytes. After were opened and plasma was pipetted and transferred into another sterile tube. Plasma was centrifuged again at 2000rpm for 10 minutes; as a result it was split into two parts: on the top, consisting of platelet-poor plasma (PPP) and at the bottom of the platelet button. Part of the PPP was discarded so that only 1ml remained in the tube along with the platelet button. This material was gently agitated to promote platelets resuspension and activated when added 0.3ml of calcium gluconate, resulting in PRP gel. Double centrifugation protocol was able to make platelet concentration 3 times higher in relation to the initial blood sample. The volume of calcium gluconate used for platelet activation was 0.3ml, and was sufficient to coagulate the sample. Coagulation time ranged from 8 to 20 minutes, with an average of 17.6 minutes. Therefore, time of blood centrifugation until to obtain PRP gel took only 40 minutes. It was concluded that PRP was successfully obtained by double centrifugation protocol, which is able to increase the platelet concentration in the sample compared with whole blood, allowing its use in surgical procedures. Furthermore, the preparation time is appropriate to obtain PRP in just 40 minutes, and calcium gluconate is able to promote the activation of platelets.

Utilização de plasma rico em plaquetas para estimulação da angiogênese em flape de padrão axial toracodorsal em coelhos (Oryctolagus cuniculus)

Resumo: Feridas de grandes extensões, com perda da viabilidade tecidual e retardo na cicatrização por segunda intenção são casos que se faz necessário o emprego de técnicas cirúrgicas reconstrutivas. O plasma rico em plaquetas (PRP) é um produto com maior concentração plaquetária, adjuvante no processo cicatricial de cirurgias reconstrutivas, auxiliando nos processos de hemostasia e estimulação da angiogênese. Dessa forma, delineou-se um estudo a fim de avaliar a eficácia do uso do gel produzido a partir do plasma rico em plaquetas (PRP) em flapes de avanço de padrão axial toracodorsal em coelhos, para avaliar a possibilidade de favorecer a integração do retalho no leito receptor. Utilizaram-se 30 coelhos da raça Nova Zelândia branco, separados em dois grupos de 15 animais, compreendendo os grupos plasma rico em plaquetas (GPRP), na qual empregou-se o gel antes da síntese da ferida cirúrgica, e controle (GC), na qual utilizou-se apenas solução fisiológica. Para obtenção do PRP, coletou-se sangue dos animais, e determinou-se a contagem plaquetária antes da preparação do gel. No início e término do experimento os animais foram pesados para posterior análise de ganho peso médio. Após o procedimento cirúrgico iniciou-se as avaliações macroscópicas no 3º, 7º e 14º dia, e avaliou-se presença ou ausência de exsudato, integridade da pele, edema, rubor e necrose. Após esta etapa, coletou-se o material da ferida cirúrgica para confecção das lâminas histológicas e posterior avaliação microscópica. Avaliou-se a proliferação vascular, presença de células mononucleares e polimorfonucleares, proliferação fibroblástica, colagenização, reepitelização e hemorragia. Os dados obtidos foram submetidos à análise estatística (Teste t Student, t emparalhado, e Kruskall Walis, sendo p<0,05). O ganho de peso médio não foi significativo entre os grupos; a concentração plaquetária da amostra final do PRP foi significativamente maior quando comparada com a inicial; exsudato e necrose foram significativamente maior no grupo controle quando comparado ao grupo PRP; proliferação vascular e reepitelização foram significativamente maior no grupo PRP, enquanto que a presença de mononucleares, polimorfonucleares, proliferação de fibroblastos, colagenização e hemorragia não foram significativas entre os grupos. Obteve-se no terceiro dia, diferença significativamente maior quanto à variável exsudato no grupo controle quando comparado ao grupo PRP, no sétimo dia exsudato e rubor foram significativamente maior no grupo controle, e ao décimo quarto dia exsudato e integridade da pele foram significativamente maior no grupo controle. Os resultados obtidos neste estudo evidenciaram que a utilização do plasma rico em plaquetas na forma de gel em cirurgia reconstrutiva foi capaz de estimular a angiogênese na ferida favorecendo o processo de cicatrização.

Chromosome damage in underground coal miners: detection by conventional cytogenetic techniques and by submitting lymphocytes of unexposed individuals to plasma from at-risk groups

Chromosome abnormalities and the mitotic index in lymphocyte cultures and micronuclei in buccal mucosa cells were investigated in a sample of underground mineral coal miners from Southern Brazil. A decreased mitotic index, an excess of micronuclei and a higher frequency of chromosome abnormalities (fragments, polyploidy and overall chromosome alterations) were observed in the miners when compared to age-paired normal controls from the same area. An alternative assay for clastogenesis in occupational exposition was tested by submitting lymphocytes from non-exposed individuals to a pool of plasmas from the exposed population. This assay proved to be very convenient, as the lymphocytes obtained from the same individuals can be used as target as well as control cells. Also, it yielded a larger number of metaphases and of successful cultures than with common lymphocyte cultures from miners. A significantly higher frequency of chromatid gaps, fragments and overall alterations were observed when lymphocytes from control subjects were exposed to miner plasma pools. Control plasma pools did not significantly induce any type of chromosome alterations in the cultures of normal subjects, thus indicating that the results are not due to the effect of the addition of plasma pools per se.

Correlations between ANP concentrations in atria, plasma and cerebral structures and sodium chloride preference in Wistar rats

We determined whether ANP (atrial natriuretic peptide) concentrations, measured by radioimmunoassay, in the ANPergic cerebral regions involved in regulation of sodium intake and excretion and pituitary gland correlated with differences in sodium preference among 40 Wistar male rats (180-220 g). Sodium preference was measured as mean spontaneous ingestion of 1.5% NaCl solution during a test period of 12 days. The relevant tissues included the olfactory bulb (OB), the posterior and anterior lobes of the pituitary gland (PP and AP, respectively), the median eminence (ME), the medial basal hypothalamus (MBH), and the region anteroventral to the third ventricle (AV3V). We also measured ANP content in the right (RA) and left atrium (LA) and plasma. The concentrations of ANP in the OB and the AP were correlated with sodium ingestion during the preceding 24 h, since an increase of ANP in these structures was associated with a reduced ingestion and vice-versa (OB: r = -0.3649, P<0.05; AP: r = -0.3291, P<0.05). Moreover, the AP exhibited a correlation between ANP concentration and mean NaCl intake (r = -0.4165, P<0.05), but this was not the case for the OB (r = 0.2422). This suggests that differences in sodium preference among individual male rats can be related to variations of AP ANP level. Earlier studies indicated that the OB is involved in the control of NaCl ingestion. Our data suggest that the OB ANP level may play a role mainly in day-to-day variations of sodium ingestion in the individual rat

Diclofenac plasma protein binding: PK-PD modelling in cardiac patients submitted to cardiopulmonary bypass

Twenty-four surgical patients of both sexes without cardiac, hepatic, renal or endocrine dysfunctions were divided into two groups: 10 cardiac surgical patients submitted to myocardial revascularization and cardiopulmonary bypass (CPB), 3 females and 7 males aged 65 ± 11 years, 74 ± 16 kg body weight, 166 ± 9 cm height and 1.80 ± 0.21 m2 body surface area (BSA), and control, 14 surgical patients not submitted to CPB, 11 female and 3 males aged 41 ± 14 years, 66 ± 14 kg body weight, 159 ± 9 cm height and 1.65 ± 0.16 m2 BSA (mean ± SD). Sodium diclofenac (1 mg/kg, im Voltaren 75® twice a day) was administered to patients in the Recovery Unit 48 h after surgery. Venous blood samples were collected during a period of 0-12 h and analgesia was measured by the visual analogue scale (VAS) during the same period. Plasma diclofenac levels were measured by high performance liquid chromatography. A two-compartment open model was applied to obtain the plasma decay curve and to estimate kinetic parameters. Plasma diclofenac protein binding decreased whereas free plasma diclofenac levels were increased five-fold in CPB patients. Data obtained for analgesia reported as the maximum effect (EMAX) were: 25% VAS (CPB) vs 10% VAS (control), P<0.05, median measured by the visual analogue scale where 100% is equivalent to the highest level of pain. To correlate the effect versus plasma diclofenac levels, the EMAX sigmoid model was applied. A prolongation of the mean residence time for maximum effect (MRTEMAX) was observed without any change in lag-time in CPB in spite of the reduced analgesia reported for these patients, during the time-dose interval. In conclusion, the extent of plasma diclofenac protein binding was influenced by CPB with clinically relevant kinetic-dynamic consequences

Does plasma ANP participate in natriuresis induced by a-MSH?

a-Melanocyte-stimulating hormone (a-MSH; 0.6 and 3 nmol) microinjected into the anteroventral region of the third ventricle (AV3V) induced a significant increase in diuresis without modifying natriuresis or kaliuresis. Intraperitoneal (ip) injection of a-MSH (3 and 9.6 nmol) induced a significant increase in urinary sodium, potassium and water excretion. Intraperitoneal (3 and 4.8 nmol) or iv (3 and 9.6 nmol) administration of a-MSH did not induce any significant changes in plasma atrial natriuretic peptide (ANP), suggesting that the natriuresis, kaliuresis and diuresis induced by the systemic action of a-MSH can be dissociated from the increase in plasma ANP. These preliminary results suggest that a-MSH may be involved in a g-MSH-independent mechanism of regulation of hydromineral metabolism

Release of plasma atrial natriuretic peptide after volume expansion is not related to pituitary-adrenal axis diurnal variation in normal subjects

The existence of a circadian rhythm of atrial natriuretic peptide (ANP) in humans is controversial. We studied the plasma ANP response to isotonic blood volume expansion in the morning and in the afternoon and its relationship with adrenocorticotropic hormone (ACTH)-cortisol diurnal variation in seven normal subjects. Basal plasma ANP level was similar in the morning (19.6 ± 2.4 pg/ml) and in the afternoon (21.8 ± 4.8 pg/ml). The ANP peak obtained with saline infusion (0.9% NaCl, 12 ml/kg) in the morning (49.4 ± 8 pg/ml) did not differ from that obtained in the afternoon (60.3 ± 10.1 pg/ml). There was no correlation between the individual mean cortisol and ACTH levels and the ANP peak obtained with saline infusion. These data indicate no diurnal variation in plasma ANP secretion induced by blood volume expansion and no relationship between plasma ANP peak and ACTH-cortisol diurnal variation

Plasma clearance and biodistribution of oxidatively modified 99mTc-ß-VLDL in rabbits

The biodistribution and removal from plasma (measured as fractional clearance rate, FCR, per hour) of native and oxidatively modified 99mtechnetium-labeled ß-very low density lipoprotein (99mTc-ß-VLDL) were investigated in hypercholesterolemic (HC) and control (C) three-month old New Zealand rabbits. The intracellular accumulation of ß-VLDL labeled with 99mTc was studied in vitro in THP-1 cells and monocyte-derived macrophages isolated from rabbits. After intravenous injection into C rabbits, copper-oxidized ß-VLDL (99mTc-ox-ß-VLDL) was cleared from the circulation faster (0.362 ± 0.070/h) than native ß-VLDL (99mTc-nat-ß-VLDL, 0.241 ± 0.070/h). In contrast, the FCR of 99mTc-ox-ß-VLDL in HC rabbits was lower (0.100 ± 0.048/h) than that of 99mTc-nat-ß-VLDL (0.163 ± 0.043/h). The hepatic uptake of radiolabeled lipoproteins was lower in HC rabbits (0.114 ± 0.071% injected dose/g tissue for 99mTc-nat-ß-VLDL and 0.116 ± 0.057% injected dose/g tissue for 99mTc-ox-ß-VLDL) than in C rabbits (0.301 ± 0.113% injected dose/g tissue for 99mTc-nat-ß-VLDL and 0.305 ± 0.149% injected dose/g tissue for 99mTc-ox-ß-VLDL). The uptake of 99mTc-nat-ß-VLDL and 99mTc-ox-ß-VLDL by atherosclerotic aorta lesions isolated from HC rabbits (99mTc-nat-ß-VLDL: 0.033 ± 0.012% injected dose/g tissue and 99mTc-ox-ß-VLDL: 0.039 ± 0.017% injected dose/g tissue) was higher in comparison to that of non-atherosclerotic aortas from C rabbits (99mTc-nat-ß-VLDL: 0.023 ± 0.010% injected dose/g tissue and 99mTc-ox-ß-VLDL: 0.019 ± 0.010% injected dose/g tissue). However, 99mTc-nat-ß-VLDL and 99mTc-ox-ß-VLDL were taken up by atherosclerotic lesions at similar rates. In vitro studies showed that both monocyte-derived macrophages isolated from rabbits and THP-1 macrophages significantly internalized more 99mTc-ox-ß-VLDL than 99mTc-nat-ß-VLDL. These results indicate that in cholesterol-fed rabbits 99mTc-ox-ß-VLDL is slowly cleared from plasma and accumulates in atherosclerotic lesions. However, although the extent of in vitro uptake of 99mTc-ox-ß-VLDL by macrophages was high, the in vivo accumulation of this radiolabeled lipoprotein by atherosclerotic lesions did not differ from that of 99mTc-nat-ß-VLDL.

Plasma cortisol levels in captive wild felines after chemical restraint

Eight Panthera onca (Po), 13 Felis concolor (Fc), 7 Felis yagouaroundi (Fy), 7 Felis tigrina (Ft) and 5 Felis pardalis (Fp) specimens from São Paulo State zoos were used. All animals were restrained with darts containing 10 mg/kg ketamine and 1 mg/kg xylazine. Venous blood samples were collected as soon as possible (within 15-20 min) and serum was frozen until the time for cortisol quantification. Cortisol was determined using a solid phase radioimmunoassay with an intra-assay coefficient of 8.51%. Data were analyzed statistically by the Kruskal-Wallis test, followed by Dunn's multiple comparisons test, and the one-sample t-test, with the level of significance set at P<0.05. Data are reported as means ± SEM. Cortisol levels differed among the captive felines: Po = 166 ± 33a, Fc = 670 ± 118b, Fy = 480 ± 83b, Ft = 237 ± 42ab, Fp = 97 ± 12a nmol/l (values followed by different superscript letters were significantly different (P<0.001)). Since most of the veterinary procedures on these species involve chemical restraint, these results show the necessity of preventive measures in order to minimize the effect of restraint stress on more susceptible species

Plasma testosterone and 11-ketotestosterone levels of male pacu Piaractus mesopotamicus (Cypriniformes, Characidae)

The levels of testosterone (T) and 11-ketotestosterone (11-KT) of the South American pacu Piaractus mesopotamicus were determined by radioimmunoassay during two stages of the reproductive cycle, i.e., resting and maturation, and the gonadosomatic index (GSI) was calculated. The highest levels of T and 11-KT were reached during the maturation stage (T = 2400 ± 56 pg/ml; 11-KT = 2300 ± 60 pg/ml) and lower levels were maintained during the resting period. The rise in androgen levels occurred with the appearance of spermatozoa in the maturation stage, when GSI was highest

A simple HPLC-fluorescence method for the measurement of R,S-sotalol in the plasma of patients with life-threatening cardiac arrhythmias

R,S-sotalol, a ß-blocker drug with class III antiarrhythmic properties, is prescribed to patients with ventricular, atrial and supraventricular arrhythmias. A simple and sensitive method based on HPLC-fluorescence is described for the quantification of R,S-sotalol racemate in 500 µl of plasma. R,S-sotalol and its internal standard (atenolol) were eluted after 5.9 and 8.5 min, respectively, from a 4-micron C18 reverse-phase column using a mobile phase consisting of 80 mM KH2PO4, pH 4.6, and acetonitrile (95:5, v/v) at a flow rate of 0.5 ml/min with detection at lex = 235 nm and lem = 310 nm, respectively. This method, validated on the basis of R,S-sotalol measurements in spiked blank plasma, presented 20 ng/ml sensitivity, 20-10,000 ng/ml linearity, and 2.9 and 4.8% intra- and interassay precision, respectively. Plasma sotalol concentrations were determined by applying this method to investigate five high-risk patients with atrial fibrillation admitted to the Emergency Service of the Medical School Hospital, who received sotalol, 160 mg po, as loading dose. Blood samples were collected from a peripheral vein at zero, 0.5, 1.0, 1.5, 2.0, 3.0, 4.0, 6.0, 8.0, 12.0 and 24.0 h after drug administration. A two-compartment open model was applied. Data obtained, expressed as mean, were: CMAX = 1230 ng/ml, TMAX = 1.8 h, AUCT = 10645 ng h-1 ml-1, Kab = 1.23 h-1, a = 0.95 h-1, ß = 0.09 h-1, t(1/2)ß = 7.8 h, ClT/F = 3.94 ml min-1 kg-1, and Vd/F = 2.53 l/kg. A good systemic availability and a fast absorption were obtained. Drug distribution was reduced to the same extent in terms of total body clearance when patients and healthy volunteers were compared, and consequently elimination half-life remained unchanged. Thus, the method described in the present study is useful for therapeutic drug monitoring purposes, pharmacokinetic investigation and pharmacokinetic-pharmacodynamic sotalol studies in patients with tachyarrhythmias.