Erding formula and bioactive markers in hyperuricemia: towards rational scientific approaches for the modernization of Traditional Chinese Medicine

Introduction: Traditional Chinese Medicine (TCM) represents one of the first holistic approaches in the world to treat and prevent disease. Herbal medicine is one of the major therapeutic remedy in TCM. It often involves multi-herb therapies instead of single herb preparations. Parallel to western medicine, hundreds of herbal formulas have been made available as finished products. Currently, the use of herbal products is popular as treatment option or to complement western medicine. Indications of the herbal formulas were established by TCM terms such as heat-clearing and/or detoxifying which lack modern pharmacological meanings. It is difficult for people without relevant background to understand such terms and their implications for treatments. Furthermore, due to the quality control issues of herbal medicines which contain multiple constituents, consumers may be confronted with the risk of using unstandardized products. Hence, in this thesis, the modernization of TCM is discussed through employing scientific pharmaceutical approaches to a traditional formula, called Erding formula (EF). The aim was to investigate if a new indication, hyperuricemia, can be assigned to a heat-clearing and detoxifying formula. Our hypothesis was: Can Erding formula be used for hyperuricemia treatment and is esculetin a bioactive marker for this new indication? Methods: A hypoxanthine and potassium oxonateinduced hyperuricemic mouse model, a xyleneinduced inflammatory mouse model, and an acetic acidinduced pain model were used to investigate EF and its constituent herbs. The quantity of esculetin was measured by high-performance liquid chromatography. The therapeutic effect of esculetin was assessed using potassium oxonate induced hyperuricemic mouse model, and esculetin and its metabolites were characterized in serum via ultra-performance liquid chromatographyquadrupole time-of-flight mass spectrometry. To develop a modern dosage form, a laboratory-scale wet bead milling approach was employed to prepare esculetin nanocrystals. The formulation was further optimized by design of experiment, and an optimized formulation was then characterized for its saturation solubility and short-term stability. Results: The study showed that EF and Viola yedoensis Makino (Viola) lowered uric acid (UA) levels, while EF and all four individual herbs had antiinflammatory and analgesic activities. These findings revealed that EF was able to treat hyperuricemia and suggested that Viola was the main herb in EF on reducing UA levels. The study showed that esculetin significantly reduced UA levels and six metabolites of esculetin were identified in serum. This confirms that esculetin was absorbed and is a suitable bioactive and quality control marker for EF in hyperuricemia treatment. An esculetin-Povacoat nanocrystal formulation with a 200 nm particle size was successfully prepared. The formulation presented up to a 1.5-fold increase in saturation solubility compared to the bulk esculetin and it was stable for 180 days. Conclusion: The studies proved that Erding formula can be used for hyperuricemia treatment with esculetin as bioactive quality control marker. As well, a new nano-sized formulation of the bioactive marker, esculetin, was created. This presented the possibility to develop an innovative nanotechnological product of the active substances derived from herbal medicine. The findings facilitated a better understanding of TCM terms and concept through mechanistic scientific experiments. This study revealed a potential pathway and an idea to modernize TCM without setting aside its unique concepts. This might increase the global acceptance of TCM products. Furthermore, the TCM concept might be useful in the development of multi-component drug products

Medicina Tradicional Chinesa (MTC) representa uma das primeiras abordagens holísticas em âmbito global para tratar e prevenir doenças. A fitoterapia consiste na principal terapia na MTC. Frequentemente, envolve terapias com múltiplas ervas em vez de preparações individuais. Paralelamente à medicina ocidental, centenas de fórmulas herbais foram disponibilizadas como produtos acabados. Atualmente, o uso de produtos fitoterápicos é popular como opção de tratamento ou para complementar a medicina ocidental. As indicações das fórmulas fitoterápicas foram estabelecidas pelos termos da MTC, tais como "limpeza pelo calor e / ou desintoxicante", que não têm significados farmacológicos modernos. É difícil para a população em geral e mesmo para profissionais sem histórico relevante na área entender tais termos e suas implicações para os tratamentos. Além disso, devido às questões de controle de qualidade dos medicamentos fitoterápicos que contêm múltiplos constituintes, os pacientes podem ser confrontados com o risco de usar produtos não padronizados. Assim, nessa tese, a modernização da MTC é discutida por meio da utilização de abordagens farmacêuticas científicas para uma fórmula tradicional, denominada fórmula de Erding (FE). O objetivo foi o de investigar se uma nova indicação, a hiperuricemia, pode ser atribuída a uma fórmula desintoxicante e de compensação de calor. Nossa hipótese foi: a fórmula de Erding pode ser usada para tratamento de hiperuricemia e a esculetina é um marcador bioativo para essa nova indicação? Foi empregado modelo de camundongo hiperuricêmico induzido por hipoxantina e oxonato de potássio, outro modelo de camundongo inflamatório induzido por xileno e, adicionalmente, modelo de dor induzida por ácido acético. Esses modelos foram usados para investigar a FE e suas ervas constituintes. A quantidade de esculetina foi determinada por cromatografia líquida de alta eficiência. O efeito terapêutico da esculetina foi avaliado utilizando modelo de camundongo hiperuricêmico induzido por oxonato de potássio, e a esculetina e seus metabólitos foram caracterizados no soro por cromatografia líquida de alto desempenho - espectrometria de massa. Para desenvolver forma farmacêutica moderna, uma abordagem de moagem em escala úmida reduzida foi empregada tendo em vista a preparação de nanocristais de esculetina. A formulação foi ainda otimizada empregado planejamento experimental. Essa fórmula foi caracterizada quanto à sua solubilidade de saturação e estabilidade a curto prazo. O estudo mostrou que a FE e a Viola yedoensis Makino (Viola) reduziram os níveis de ácido úrico (AU), enquanto a FE e as quatro plantas individuais apresentaram atividades antiinflamatória e analgésica. Esses resultados revelaram que a FE foi capaz de tratar a hiperuricemia e sugeriu que a viola foi a principal erva da FE na redução dos níveis de AU. O estudo mostrou também que a esculetina reduziu significativamente os níveis de AU e os seis metabólitos da esculetina foram identificados no soro. Tal resultado confirma que a esculetina foi absorvida e pode ser usada como marcador de controle bioativo e de qualidade para FE, no tratamento da hiperuricemia. A formulação de nanocristais de esculetin-povacoat® apresentou tamanho de partícula de 200 nm. A formulação apresentou aumento de 1,5 vezes na solubilidade de saturação em comparação com a esculetina em escala micrométrica e manteve-se estável durante 180 dias. Os estudos comprovaram que a fórmula de Erding pode ser utilizada no tratamento da hiperuricemia empregando a esculetina como marcador bioativo de controle de qualidade. Além disso, foi desenvolvida formulação inovadora, em escala nanométrica, do marcador bioativo, a esculetina. Esse resultado permitiu desenvolver produto com base nanotecnológica das substâncias ativas derivadas do fitoterápico, assim comol permitiram melhor compreensão dos termos e dos conceitos da MTC por meio de experimentos científicos mecanicistas. Esse estudo revelou potencial para a modernização da MTC sem excluir seus conceitos únicos. Isso pode aumentar a aceitação global dos produtos MTC. Além disso, o conceito de MTC pode ser útil no desenvolvimento de medicamentos de múltiplos componentes

Matrix effects of the hydroethanolic extract and the butanol fraction of calyces from Physalis peruviana L. on the biopharmaceutics classification of rutin.

OBJECTIVES: The Biopharmaceutics Classification System (BCS) categorizes active pharmaceutical ingredients according to their solubility and permeability properties, which are susceptible to matrix or formulation effects. The aim of this research was to evaluate the matrix effects of a hydroethanolic extract of calyces from Physalis peruviana L. (HEE) and its butanol fraction (BF), on the biopharmaceutics classification of their major compound, quercetin-3-O-rutinoside (rutin, RU). METHODS: Rutin was quantified by HPLC-UV, and Caco-2 cell monolayer transport studies were performed to obtain the apparent permeability values (Papp ). Aqueous solubility was determined at pH 6.8 and 7.4. KEY FINDINGS: The Papp values followed this order: BF > HEE > RU (1.77 ± 0.02 > 1.53 ± 0.07 > 0.90 ± 0.03 × 10-5  cm/s). The lowest solubility values followed this order: HEE > RU > BF (2.988 ± 0.07 > 0.205 ± 0.002 > 0.189 ± 0.005 mg/ml). CONCLUSIONS: According to these results, rutin could be classified as BCS classes III (high solubility/low permeability) and IV (low solubility/low permeability), depending on the plant matrix. Further work needs to be done in order to establish how apply the BCS for research and development of new botanical drugs or for bioequivalence purposes.

[Research and development thought on biopharmaceutics classification system of Chinese materia medica].

The biopharmaceutics classification system( BCS) is a scientific framework or method for classifying drugs based on drug solubility and permeability,which can be used to provide drug bioavailability-absorption correlation analysis. Based on the characteristics of multi-component and multi-target of traditional Chinese medicine( TCM) as well as the concept,method and technology of BCS,the research group proposed biopharmaceutics classification system of Chinese materia medica( CMMBCS) and carried out research and data accumulation of classical prescriptions. Based on the previous research results,further development ideas under the CMMBCS concept and framework were further proposed in this study. In the course of research,the influence of the intermediate links of the complex interactions of the multi-component environment was omitted,and the component absorption studies on the main clinical effects of prescription ingredients were directly concerned,or the components and data were reversely extracted from the aspects of metabolism,pharmacodynamic pathways and absorption principles. Studies were conducted from two aspects( single component and compound prescription) to comprehensively evaluate the absorption properties of TCM compound. In the research path,the different ways in which Chinese medicine could exert its efficacy were fully considered,and CMMBCS classification and establishment rules were clarified mainly by focusing on the absorption pathway into the blood. Specifically,the network pharmacology and molecular docking technology were used to screen the compound index components of TCM; the absorption rules were studied by the physiologically based pharmacokinetic models and the absorption parameters of CMMBCS were calculated by reverse reasoning. Then the CMMBCS classification of TCM prescription was corrected by studying the efficacy or absorption pathway. In this paper,the theoretical framework and research methodology of CMMBCS were systematically improved based on the establishment of CMMBCS basic theory,the supplementary of drug-oriented research ideas and the application of modern mature Chinese medicine methodology.

[Research on attributes of biopharmaceutics classification system for Chinese materia medica of baicalein in Gegen Qinlian Decoction environment].

For the effects of multi-component environment on the solubility and permeability of single components,and the problems of biopharmaceutical attribute classification of single components in the compound prescriptions environment,baicalein was used as the research object in this study to investigate the biopharmaceutic attributes of single-component and their traditional Chinese medicine( TCM) biopharmaceutic attributes in the multi-component environment of Gegen Qilian Decoction. Shaking flask method,intrinsic dissolution rate test and HPLC were used to determine solubility of baicalein. Markers specified by FDA were utilized as permeable boundary reference materials to verify the applicability of the single-pass intestinal perfusion method( SPIP),and the quantitative research on the permeability of baicalein was also conducted. It is concluded that baicalein could be categorized as BCS-Ⅱ drug based on its low solubility and high intestinal permeability values,and it may be categorized into CMMBCS-I in the multi-component environment of Gegen Qilian Decoction due to its poor solubility but enhanced solubility and permeability in compound environment. This study could provide verification ideas for clinical determination of the best human oral dose of baicalein,and provide the data basis for the study of biopharmaceutics classification system of Chinese materia medica( CMMBCS).

[Study on biopharmaceutics classification system of Chinese materia medica for Gegen Qinlians Tablets based on anti-inflammatory activity].

The research on biopharmaceutics classification system of Chinese materia medica( CMMBCS) should be finally implemented to the holistic research level of traditional Chinese medicine compounds,while the overall biopharmaceutical properties of traditional Chinese medicine compounds are not only the sum of solubility and permeability of each component. In this study,Gegen Qinlian Tablets was used as the research object,and the contents of 12 representative components,i.e. puerarin,daidzin,baicalin,daidzein,wogonoside,baicalein,wogonin,glycyrrhizic acid,coptisine hydrochloride,epiberberine,berberine hydrochloride and palmatine hydrochloride,were simultaneously determined by HPLC to obtain the mass weight of each component. The in vitro lipopolysaccharide( LPS)-induced RAW264. 7 cells inflammation model was established to investigate the anti-inflammatory effects of 12 representative components and obtain the efficacy weight of each component. In order to obtain the number of doses and effective permeability coefficient which can represent the overall biopharmaceutical properties of Gegen Qinlian Tablets,mass weight was combined with efficacy weight to integrate the solubility and permeability data of each component determined by typical shake flask method and in situ single pass intestinal perfusion model respectively. The results indicated that Gegen Qinlian Tablets should be categorized Ⅳ drug of the CMMBCS with low solubility and low permeability.

Application of a Refined Developability Classification System.

In 2010, the Developability Classification System was proposed as an extension of the Biopharmaceutics Classification System to align the classification system with the need for early evaluation of drug candidates according to their developability as oral formulations. Recent work on the Developability Classification System has resulted in the refined developability classification system (rDCS), consisting of standard investigations to estimate drug candidate solubility and permeability and offering customized investigations that are triggered when there is a potential for supersaturation/precipitation (e.g., salts of acids, weak bases) or to investigate permeation versus dissolution-limited absorption. In the present study, the rDCS concept was successfully applied to 6 marketed compounds (aciclovir, albendazole, danazol, dantrolene, dipyridamole, and piroxicam), for which there is a rich database of information. Furthermore, the rDCS was applied to 20 pipeline compounds from past and current research projects at Bayer AG. The rDCS was able to predict the results in humans correctly in 80% of cases. Overall, the results suggest that the rDCS is a highly useful tool for estimating the in vivo behavior of new drug candidates.

Comparative Oral Drug Classification Systems: Acetazolamide, Azithromycin, Clopidogrel, and Efavirenz Case Studies.

Biopharmaceutics classification systems based on the properties of solubility and permeability or the extension of metabolism are very important tools in the early stages of the development and regulatory stages of new products. However, until now, there was no clear understanding between the interplay among these classification systems. Therefore, the main objective of this work was to make a comparison of concepts of BCS and BDDCS to understand what are the key factors that allow for the integration of these biopharmaceutics classification systems. Also, the suitability of an in situ single-pass intestinal perfusion assay in rats (SPIP) development was assessed by us to determine the limit between high and low permeability following what the FDA BCS guidance suggests. An excellent correlation was found between the values of permeability obtained by applying SPIP assays and the extensions of the metabolism of the set of compounds studied in this work, with the exception of three compounds that showed disparity between their permeability coefficients ( Peff), obtained herein by SPIP, and their metabolism (acetazolamide, azithromycin, and efavirenz). Discrepancies allowed us to elucidate the interrelationship between BCS and BDDCS.

[Biopharmaceutics classification system of Chinese materia medica of berberine in Gegen Qinlian decoction].

One of the top-level researches of biopharmaceutics classification system of Chinese materia medica (CMMBCS) is the study on single component in compound Chinese medicine. The medicines shall be classified according to its solubility and intestinal permeability, as well as the ascending degree in multicomponent environment. Based on above, we chose berberine as the main object to explore the change rules of its solubility and intestinal permeability in Gegen Qinlian decoction. Shaking flask-HPLC was used to detect the solubility changes of berberine in compounds. The qualitative investigation of berberine in intestinal absorption was measured by everted gut sac, and the quantitative research of berberine in intestinal absorption was measured by single-pass intestinal perfusion experiment, while the qualitative and quantitative research of berberine absorption into blood was measured by in intestinal perfusion with venous sampling experiment.

Evaluation of DILI Predictive Hypotheses in Early Drug Development.

Drug-induced liver injury (DILI) is a leading cause of drug failure in clinical trials and a major reason for drug withdrawals. DILI has been shown to be dependent on both daily dose and extent of hepatic metabolism. Yet, early in drug development daily dose is unknown. Here, we perform a comprehensive analysis of the published hypotheses that attempt to predict DILI, including a new analysis of the Biopharmaceutics Drug Disposition Classification System (BDDCS) in evaluating the severity of DILI warnings in drug labels approved by the FDA and the withdrawal status due to adverse drug reactions (ADRs). Our analysis confirms that higher doses ≥50 mg/day lead to increased DILI potential, but this property alone is not sufficient to predict the DILI potential. We evaluate prior attempts to categorize DILI such as Rule of 2, BSEP inhibition, and measures of key mechanisms of toxicity compared to BDDCS classification. Our results show that BDDCS Class 2 drugs exhibit the highest DILI severity and that all of the published methodologies evaluated here, except when daily dose is known, do not yield markedly better predictions than BDDCS. The assertion that extensive metabolized compounds are at higher risk of developing DILI is confirmed but can be enhanced by differentiating BDDCS Class 2 from Class 1 drugs. We do not propose that the BDDCS classification, which does not require knowledge of the clinical dose, is sufficiently predictive/accurate of DILI potential for new molecular entities but suggest that comparison of proposed DILI prediction methodologies with BDDCS classification is a useful tool to evaluate the potential reliability of newly proposed algorithms. CONCLUSION: The most successful approaches to predict DILI potential all include a measure of dose, yet there is a quantifiable uncertainty associated with the predicted dose early in drug development. Here, we compare the possibility of predicting DILI potential using the BDDCS classification versus previously published methods and note that many hypothesized predictive DILI metrics do no better than just avoiding BDDCS Class 2 drugs.

[Study on property of biopharmaceutics classification system of berberine in Huanglian decoction].

The study of single component in the multicomponent environment is one of the basic researches for biopharmaceutics classification system of Chinese materia medica (CMMBCS). That is to say, the classification research shall be based on the respective lift of solubility and permeability in the multicomponent environment, besides solubility and intestinal permeability of the single component. We chose berberine as the main research object to investigate the changes of its solubility and intestinal permeability in Huanglian decoction. Shake-flask and HPLC were used to detect the solubility of berberine in different pH buffer solutions and different concentrations of Huanglian decoction. In situ single-pass intestinal perfusion (SPIP) and intestinal perfusion with venous sampling (IPVS) were carried out to study berberine's intestinal absorption and absorption into blood, respectively.

[Biopharmaceutics classification and absorption mechanisms primary study on four kinds of flavonoids].

The solubility and permeability on four kinds of flavonoids (kaempferol, hesperidin, apigenin, genistein) were test according to the theory of biopharmaceutics classification system (BCS), and their absorption mechanism. The solubility was investigated by the method in determination of solubility of "Chinese Pharmacopoeia 2010". To detect appearance permeability of compounds mentioned above, the appropriate concentrations were selected by the MTT method in cell transfer experiments in Caco-2 cell model, which established by in vitro cell culture method. Therefore, these compounds were classified with BCS according to solubility and permeability. In addition, to explore absorption mechanisms, the experiments in three different concentrations of compounds in high, medium and low in bidirectional transformation methods in Caco-2 cell model contacted. The study indicated that all of kaempferol, hesperidin, apigenin, genistein have the characteristics in low solubility and high permeability, which belong to BCSⅡ, and the absorption mechanism of kaempferol was active transportation. Whereas, hesperidin, apigenin, genistein were passive transportation. In this study, it carried out initial explorations on establishment of determination for solubility and permeability in flavonoids, and provided theoretical reference for further research on BCS in traditional Chinese medicine.

[Biotherapies, immunotherapies, targeted therapies, biopharmaceuticals which word should be used?]. / Biothérapies, immunothérapies, thérapies ciblées, biomédicaments - De quoi faut-il parler ?

The ability to accurately describe and name medical advances is a prerequisite to foster public debates with scientists and physicians, and favour faith over fear among patients and citizens. Therapeutic antibodies are a good example of a medical breakthrough which has met with considerable clinical success, and which terminology has changed over the years. If the appellation serotherapy was appropriate a century ago, it has become obsolete. Recent names such as biotherapy, immunotherapy, targeted therapy, biopharmaceuticals have been introduced and are now commonly used, each of those can apply to therapeutic antibodies. It is thus interesting to question the real meaning of these different appellations. Our goal in this manuscript is to analyse the genesis of these terms but also to suggest how to simplify the terminology: biotherapy or targeted therapy need to be eliminated, as well as immunotherapy when communicating with non scientific public. It is recommended to favour the term biopharmaceuticals (biomédicaments in French), which clearly indicates the origin of these molecules, intermediate between chemical drugs and living biologics, whose borders need to be accurately defined also.

The use of biopharmaceutic classification of drugs in drug discovery and development: current status and future extension.

Bioavailability (BA) and bioequivalence (BE) play a central role in pharmaceutical product development and BE studies are presently being conducted for New Drug Applications (NDAs) of new compounds, in supplementary NDAs for new medical indications and product line extensions, in Abbreviated New Drug Applications (ANDAs) of generic products and in applications for scale-up and post-approval changes. The Biopharmaceutics Classification System (BCS) has been developed to provide a scientific approach for classifying drug compounds based on solubility as related to dose and intestinal permeability in combination with the dissolution properties of the oral immediaterelease (IR) dosage form. The aim of the BCS is to provide a regulatory tool for replacing certain BE studies by accurate in-vitro dissolution tests. The aim of this review is to present the status of the BCS and discuss its future application in pharmaceutical product development. The future application of the BCS is most likely increasingly important when the present framework gains increased recognition, which will probably be the case if the BCS borders for certain class II and III drugs are extended. The future revision of the BCS guidelines by the regulatory agencies in communication with academic and industrial scientists is exciting and will hopefully result in an increased applicability in drug development. Finally, we emphasize the great use of the BCS as a simple tool in early drug development to determine the rate-limiting step in the oral absorption process, which has facilitated the information between different experts involved in the overall drug development process. This increased awareness of a proper biopharmaceutical characterization of new drugs may in the future result in drug molecules with a sufficiently high permeability, solubility and dissolution rate, and that will automatically increase the importance of the BCS as a regulatory tool over time.

Development of a more rapid, reduced serum culture system for Caco-2 monolayers and application to the biopharmaceutics classification system.

The objectives were: (1) to develop a more rapid, reduced serum culture system for Caco-2 monolayers, relative to the traditional 21-day, 10% fetal bovine serum (FBS) system; and (2) to determine the biopharmaceutical drug classification of an oral therapeutic agent using this new system. Caco-2 cells were grown in the six well format on polycarbonate filters, in medium containing 2% iron supplemented calf serum (sCS) and a combination of growth factors and hormones. After 4 days in culture, permeabilities of three marker compounds (metoprolol, mannitol, and taurocholate) across monolayers were determined, and compared to permeabilities from the traditional 21-day, 10% FBS system, using cells at similar passage number. Cell morphology, degree of cell differentiation, and the presence of two efflux pumps were assessed. The 2% sCS model was also used to classify the permeability of an oral therapeutic agent as high or low. No difference in permeability was observed for metoprolol transport (P = 0.38) between the two culture methods, and the values obtained were independent of passage number of the cells. Mannitol permeability was about 2-fold higher from the 2% sCS system, as compared to the 10% FBS system. Taurocholate permeability was low indicating the 2% sCS culture at 4 days lacked this particular active transporter capability. Electron micrographs of cells grown in the 2% sCS system at 4 days revealed the presence of microvilli and tight junctions. P-glycoprotein and an efflux pump for furosemide were functionally present. The 2% sCS system indicated the oral therapeutic agent as highly permeable, which agreed with the 10% FBS system. This new system provides a rapid, accurate, and economical option for passive permeability determination, and appears to be applicable to the proposed Biopharmaceutics Classification System (BCS).