Dúvidas denucci@gdenucci.com Arquivo Estudos de Eficácia com Animais Sadios (biodisponibilidade e bioequivalência) Site www.gdenucci.com

Administração endovenosa de 200mg em bôlus - concentração plasmática vs tempo após administração Tempo (h) 1.0 2.0 4.0 6.0 8.0 10.0 12.0 18.0 24.0 36.0 48.0 mg/L 15.800 12.600 8.000 5.000 3.200 2.000 1.260 0.320 0.080 0.000

Administração endovenosa 10 20 30 40 50 60 Tempo (h) Concentração (mg/L)

Cálculo da meia-vida 100 10 Concentração (mg/L) 5 t½ 1 10 20 Tempo (h)

Meia-Vida Primeira meia-vida – 50% do medicamento é eliminado Segunda meia-vida – 75% do medicamento é eliminado Terceira meia-vida – 87.5% do medicamento é eliminado Após cinco meias-vidas, praticamente 100% do medicamento é eliminado

Concentration Mean Diazepam Valium 300 Test formulation 200 Concentration (ng/mL) 100 4 8 12 16 20 24 48 96 144 192 240 Time (h)

Concentration Mean Nordiazepam Valium 4 8 12 16 20 24 10 30 40 Test formulation Concentration (ng/mL) 48 96 144 192 240 Time (h)

Administração de uma cápsula de 150 mg de Venlafaxina a um voluntário sadio – curva de concentração em função do tempo

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – curva de concentração em função do tempo 7 6 5 4 Concentração Plasmática (mg/L) 3 2 1 1 2 3 4 5 6 7 8 9 10 11 12 Tempo (h)

Administração de um comprimido de 5 mg de Amlodipina a um voluntário sadio – curva de concentração em função do tempo 7.5 5.0 Concentração Plasmática (µg/L) 2.5 0.0 2 4 6 8 10 12 14 24 48 72 96 120 144 Tempo (h)

Concentração Plasmática Amoxicilina 110 7000 Venlafaxina 100 6000 90 80 5000 70 4000 Concentração Plasmática (ug/L) 60 Concentração Plasmática (ug/mL) 50 3000 40 30 2000 20 1000 10 24 48 Tempo (h)

Concentração Plasmática Anlodipina 110 7 100 Venlafaxina 6 90 80 5 70 4 Concentração Plasmática 60 (ug/L) (µg/L) 50 3 Concentração Plasmática 40 30 2 20 1 10 24 48 72 96 120 144 Tempo (h)

Administração de uma cápsula de 150 mg de Venlafaxina a um voluntário sadio – curva de concentração em função do tempo Cmax

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – curva de concentração em função do tempo Cmax

Administração de um comprimido de 5 mg de Amlodipina a um voluntário sadio – curva de concentração em função do tempo Cmax

Administração de uma cápsula de 150 mg de Venlafaxina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de um comprimido de 5 mg de Amlodipina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de uma cápsula de 150 mg de Venlafaxina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de um comprimido de 5 mg de Amlodipina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – curva de concentração em função do tempo Cmax Tmax

Administração de uma cápsula de 150 mg de Venlafaxina a um voluntário sadio – área sob a curva ASC

Administração de um comprimido de 500 mg de Amoxicilina a um voluntário sadio – área sob a curva ASC

Administração de um comprimido de 5 mg de Amlodipina a um voluntário sadio – área sob a curva 2 4 6 8 10 12 14 0.0 2.5 5.0 7.5 24 48 72 96 120 144 Tempo (h) Concentração Plasmática (µg/L) ASC

Biodisponibilidade A fração da droga não modificada que atinge a circulação sistêmica independente da via de administração Biodisponibilidade absoluta – via oral comparada à via endovenosa Biodisponibilidade relativa – comparação de duas outras vias

Biodisponibilidade (F) ASC VO F = ASC EV

Biodisponibilidade relativa ASC VO teste Frel = ASC VO referência

Administração Oral Luz Intestinal Parede Intestinal Fígado Veia Porta Local de Medida Metabolismo Metabolismo Fezes

Medicamento Genérico Apresenta a mesma eficácia que o medicamento de referência, visto ter sido avaliado através de equivalência farmacêutica e/ou bioequivalência.

Equivalência Farmacêutica São testes in vitro que avaliam: Teor de princípio ativo Características físicas da formulação Perfil de dissolução em meios aquosos

Medicamento Genérico Apresenta a mesma eficácia que o medicamento de referência, visto ter sido avaliado através de equivalência farmacêutica e/ou bioequivalência.

Bioequivalência São testes realizados em humanos para avaliar a biodisponibilidade de 2 ou mais formulações

Premissa Básica da Bioequivalência Se dois medicamentos são bioequivalentes, eles apresentam a mesma eficácia terapêutica, e portanto são intercambiáveis.

Bioequivalência Em uma ocasião o voluntário ou o paciente toma uma formulação (medicamento referência por exemplo) e em outra ocasião ele toma a outra formulação (formulação teste)

Determinação da Bioequivalência Concentração da droga ativa ou metabólito(s) em fluidos biológicos em função do tempo. Excreção urinária da droga ativa ou metabólito(s) em função do tempo. Qualquer efeito farmacológico agudo apropriado.

Determinação da Bioequivalência Em 99.9% dos casos, a avaliação da bioequivalência é feita através da determinação da concentração da droga ativa ou metabólito(s) em fluidos biológicos (plasma) em função do tempo. Este estudo é feito em voluntários sadios de ambos os sexos, tratando-se de estudo clínico aberto, cruzado de duas fases e aleatorizado.

Parâmetros para estabelecimento de bioequivalência ASC - extensão da absorção Cmax - pico da absorção

Concentração plasmática da droga Parâmetros farmacocinéticos (após administração oral de um medicamento) 8 6 4 2 Concentração plasmática da droga ASC 0-12h 0 3 6 9 12 Horas

Concentração plasmática da droga Parâmetros farmacocinéticos (após administração oral de um medicamento) 8 6 4 2 Cmax Concentração plasmática da droga tmax 0 3 6 9 12 Horas

Análise estatística para bioequivalência Razões da AUC e Cmax log-transformadas Análise Paramétrica ou não-paramétrica

Análise estatística para bioequivalência Calcular intervalos de confiança (90%). Os intervalos de confiança devem variar entre 0.8-1.25 (corresponde a + 20%)

Concentração plasmática de sertralina em função do tempo (média de 24 vols) 30 Zoloft (medicamento referência) Tolrest (medicamento similar) 20 ng/mL 10 2 4 6 8 10 12 14 16 18 24 48 72 96 Horas

Análise estatística de bioequivalência TOL/ZOL Média Geométrica 90% IC AUC(0-inf) 94.6 89.3 – 100.3 Cmax 99.9 91.73 – 108.8

Objetivo da bioequivalência Assegurar o mesmo efeito terapêutico ou assegurar a similaridade das formulações farmacêuticas?

Concentration Mean

Statistical Analysis for BE 90% CI Parameters Clarithromycin OH-Clarithromycin AUC % ratio 91.1 - 103.7 94.2 - 108.1 (0 - last) AUC % ratio 91.0 - 103.4 93.9 - 107.2 (0 - inf) C % ratio 87.5 - 108.7 92.3 - 116.8 max

Statistical Analysis for BE 90% CI Parameters Diazepam Nordiazepam AUC % ratio 89.00 - 106.65 88.70 - 108.91 (0 - last) C % ratio 50.27 - 72.42 88.28 - 107.52 max

Statistical Analysis for BE 90% CI Parameters Risperidone OH-Risperidone AUC % ratio 78.60 % - 98.28 % 94.26 % - 105.94 % (0 - last) AUC % ratio 79.30 % - 98.95 % 92.54 % - 105.08 % (0 - inf) C % ratio 71.55 % - 92.91 % 86.37 % - 99.09 % max

J Vet Pharmacol Ther. 2005 Oct;28(5):419-23. Pharmacokinetics of two once-daily parenteral cephalexin formulations in dogs. Rebuelto M, Montoya L, Kreil V, Ambros L, Waxman S, Albarellos G, Hallu R. Farmacologia, Departamento de Fisiopatologia y Etiopatogenia, Facultad de Ciencias Veterinarias, Universidad de Buenos Aires, Buenos Aires, Argentina. rebuelto@fvet.uba.ar The aims of this study were to describe and compare the pharmacokinetic profiles and T(>MIC90) of two commercially available once-daily recommended cephalexin formulations in healthy adult dogs administered by the intramuscular (i.m.) route. Six beagle dogs received a 10 mg/kg dose of an 18% parenteral suspension of cephalexin of laboratory A (formulation A) and laboratory B (formulation B) 3 weeks apart. Blood samples were collected in predetermined times after drug administration. The main pharmacokinetic parameters were (mean +/- SD): AUC((0-infinity)), 72.44 +/- 15.9 and 60.83 +/- 13.2 microg.h/mL; C(max), 10.11 +/- 1.5 and 8.50 +/- 1.9 microg/mL; terminal half-life, 3.56 +/- 1.5 and 2.57 +/- 0.72 h and MRT((0-infinity)), 5.86 +/- 1.5 and 5.36 +/- 1.2 h for formulations A and B, respectively. T(>MIC90) was 63.1 +/- 14.7 and 62.1 +/- 14.7% of the dosing interval for formulations A and B, respectively. Median (range) for t(max) was 2.0 (2.0-3.0) h and 3.0 (2.0-4.0) for formulations A and B, respectively. Geometric mean ratios of natural log-transformed AUC((0-infinity)) and C(max) and their 90% confidence intervals (CI) were 0.84 (0.72-0.98) and 0.83 (0.64-1.07), respectively. The plasma profiles of cephalexin following the administration of both formulations were similar. No statistical differences between pharmacokinetic parameters or T(>MIC90) were observed, however, bioequivalence between both formulations could not be demonstrated, as lower 90% CI failed to fell within the selected range of 80-125% for bioequivalence.

Biowaivers for oral immediate-release products: implications of linear pharmacokinetics. Faassen F, Vromans H. Department of Pharmaceutics, NV Organon, Oss, The Netherlands. fried.faassen@organon.com Bioequivalence of drug formulations plays an important role in drug development. Recently, the Biopharmaceutical Classification System (BCS) has been implemented for the purpose of waiving bioequivalence studies on the basis of the solubility and gastrointestinal permeability of drug substance. Using the rationale of the BCS, it can be argued that biowaivers can, however, also be granted on the basis of standard pharmacokinetic data. If a drug exhibits dose-linear pharmacokinetics and a sufficiently fast dissolution profile, it can be concluded that this drug appears to pose no problem with respect to absorption. It should be noted that a change of an immediate-release tablet formulation can only lead to a deviating rate and/or extent of absorption when release of the drug from the formulation is altered. Logically, the dissolution profiles of the different formulations should be equal to guarantee bioequivalency. Thus, both BCS and the alternative linear pharmacokinetics approach require an evaluation of dissolution profiles. The justification of BCS is found in the permeability classification of the compound, while those of the linear pharmacokinetics lie in the apparent lack of a permeability problem. For example, in this context P-glycoprotein-transported drugs form an interesting class of compounds, which may be treated likewise when complying to the aforementioned requirements. Furthermore, poorly soluble compounds may be less troublesome than expected. It is shown that linear kinetics can be explained by the solubilising activity of, for example, bile salts. In this instance, linear pharmacokinetics shows that elevated doses do not appear to exhibit a limiting role on the dissolution. Hence, a change in formulation without any effect on the dissolution profile is not expected to cause a change in availability. It is clear that the formulations to be compared should not contain excipients that display an effect on (presystemic) drug metabolism.

J Pharm Biomed Anal. 2005 Feb 7;37(1):187-93. High-performance liquid chromatography electrospray ionization mass spectrometry determination of tulobuterol in rabbit's plasma. Xu F, Zhang Z, Tian Y, Jiao H, Liang J, Gong G. Center for Instrumental Analysis, China Pharmaceutical University, Nanjing 210009, PR China. A sensitive and specific liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) method has been developed and validated for the identification and quantification of tulobuterol in rabbits' plasma. After the addition of clenbuterol-HCl, the internal standard (IS) and 1.0 M sodium hydroxide solution, plasma samples were extracted using a solvent mixture comprised of 5% isopropanol in n-hexane. The compounds were separated on a prepacked Lichrospher CN (5 microm, 150 mm x 2.0 mm) column using a mixture of methanol-water (10 mM CH3COONH4, pH 4.0) as mobile phase. A Shimadzu LCMS-2010A mass spectrometer connected to a Shimadzu high performance liquid chromatograph (HPLC) was used to develop and validate the method. The method has shown to be sensitive and specific by testing six different blank plasma batches. Linearity was established for the range of concentrations 0.50-40.0 ng/mL with a coefficient of determination (r) of 0.9998. The intra-day precision was better than 15%. The lower limit of quantification (LLOQ) was identifiable and reproducible at 0.50 ng/mL. The proposed method enables the unambiguous identification and quantification of tulobuterol for pharmacokinetic, bioavailability or bioequivalence studies.

Evaluation of the bioequivalence of two formulations of deltamethrin for treatment of sheep with psoroptic mange. Cadiergues MC, Laguerre C, Roques M, Franc M. UMR 181 Physiopatholog ie et Toxicologie Experimentales INRA/ENVT, Ecole Nationale Veterinaire de Toulouse, 23, chemin des Capelles 31076 Toulouse 3, France. OBJECTIVE: To evaluate efficacy of 2 de ltamethrin emulsifiable concentrates that differed on the basis of vehicle (methyl glycol acetate [AMG] or 2-propylene glycol 1-methyl ether acetate [AMP]) for the treatment of sheep with mange. ANIMALS: 30 ewes between 11 months and 7 years old that weighed 16 to 71 kg and were naturally infested with Psoroptes ovis. PROCEDURE: Sheep were randomly allocated into 3 groups (13 sheep in group AMP, 13 sheep in group AMG, and 4 negative-control sheep). Each sheep was dipped twice (10-day interval between dippings) in the assigned formulation. Assessment of efficacy was performed on days 3, 7, 14, 21, 28, 35, 42, 49, 56, and 63 after the first dipping. Efficacy was assessed by determining the number of eggs or live mites on those days, as well as regrowth of wool at the end of the study. RESULTS: Psoroptic mange infestation was maintained in the 4 control sheep throughout the study. We did not detect live Psoroptes mites in scrapings after day 7 (AMP group) or after day 14 (AMG group). No parasites were seen after day 14 in either treatment group. Therefore, efficacy was 100% for both treatment groups from days 14 to 63. CONCLUSIONS AND CLINICAL RELEVANCE: The 2 formulations of deltamethrin were equally able to eradicate Psoroptes infestation of sheep after 2 dippings performed in accordance with the label recommendations.

Lack of bioequivalence of two oxytetracycline formulations in the rabbit. Chong W, Kim YJ, Kim SD, Han SK, Ryu PD. Department of Pharmacology, College of Veterinary Medicine and School of Agricultural Biotechnolog, Seoul National University, Suwon, 441-744, Korea. Oxytetracycline (OTC) has been used for over 40 years in veterinary medical field. Various forms of oxytetracycline preparations have been marketed, but little information is available on the bioequivalence of OTC preparations. This study was conducted to evaluate the bioequivalence of two OTC powder preparations available in Korea. Fourteen rabbits were randomly allocated into two groups. During the first period, a dose (200 mg/kg) of reference product was orally administered to the rabbits in Group A and test product to those in Group B. After 7-day washout period the reference and test products were given in group B and A, respectively. Blood samples were drawn at 17 points during 48 hours after administration and plasma OTC concentrations were measured by using HPLC. The solution concentrations of OTC dissolved from two products were not significantly different in the dissolution test. The mean area under the curve (AUC0- infinity ) and peak plasma concentration (C(max)) values for test and reference OTCs were 7.22 +/- 3.90 and 11.04 +/- 7.37 microg h/ml, 1.11 +/- 0.65 and 1.85 +/- 1.15 microg/ml, respectively. The relative bioavailability and C(max) of test product to those of reference product was 65.4% and 60.0%, respectively. The ranges of AUC and C(max) of test drug compared to those of reference drug under 90% confidence limits were 27 104% and 28 91.5%, respectively. The results of statistical analysis indicate that the two pivotal pharmacokinetic parameters, AUC and C(max) of test product are not within the 20% of those of the reference, suggesting that the test OTC is not bioequivalent to the reference OTC.

The steady-state pharmacokinetics and bioequivalence of carprofen administered orally and subcutaneously in dogs. Clark TP, Chieffo C, Huhn JC, Nimz EL, Wang C, Boy MG. Veterinary Medicine Pharmaceuticals Clinical Development, Pfizer Global Research and Development, Pfizer, Inc., Groton, CT 06340, USA. clarktp@lilly.com Eighteen male Beagle dogs were randomized to oral (p.o.) or subcutaneous (s.c.) carprofen administration in a two-sequence, two-period crossover design with a 10-day washout between periods. Twenty-five milligrams of carprofen was administered p.o. or s.c. every 12 h for 7 days. Plasma concentrations of carprofen collected after the first and last treatments were determined by high-performance liquid chromatography. Carprofen concentration data were natural log transformed and geometric means were calculated for maximum plasma concentration (Cmax) and area under the plasma concentration-time curve (AUC0--12) following the first dose and Cmax and AUC0--12 following administration of the last dose. Formulations were considered bioequivalent if the 90% confidence interval (CI) of the mean difference for each variable between formulations were within -20% and 25% of the oral formulation. The mean Cmax and AUC0--12 were 16.9 microg/mL and 73.1 microg. h/mL, respectively, following a single oral dose and 8.0 microg/mL and 64.3 microg x h/mL, respectively, following a single s.c. injection. The 90% CI for Cmax (-56.8 to -48.7%) was outside of the bioequivalence criteria whereas the 90% CI for AUC0--12 (-16.3 to -7.5%) was within the bioequivalence criteria. At steady-state, the mean Cmax and AUC0--12 were 18.7 microg/mL and 101.9 microg x h/mL, respectively, following p.o. administration and 14.7 microg/mL and 111.0 microg x h/mL, respectively, following s.c. injection. The 90% CI was outside the bioequivalence criteria for Cmax (-30.8 to -10.8) but within the bioequivalence criteria for AUC0--12 (2.3-15.9%). The results of this study indicate that peak plasma concentrations of carprofen differ when administered p.o. and s.c., but that total drug exposure following a single dose and at steady-state are bioequivalent.