RESUMO
A stereogenic center, placed at an exocyclic location next to a chiral carbon in a ring to which it is attached, is a ubiquitous structural motif seen in many bioactive natural products, including di- and triterpenes and steroids. Installation of these centers has been a long-standing problem in organic chemistry. Few classes of compounds illustrate this problem better than serrulatanes and amphilectanes, which carry multiple methyl-bearing exocyclic chiral centers. Nickel-catalyzed asymmetric hydrovinylation (AHV) of vinylarenes and 1,3-dienes such as 1-vinylcycloalkenes provides an exceptionally facile way of introducing these chiral centers. This Article documents our efforts to demonstrate the generality of AHV to access not only the natural products but also their various diastereoisomeric derivatives. Key to success here is the availability of highly tunable phosphoramidite Ni(II) complexes useful for overcoming the inherent selectivity of the chiral intermediates. The yields for hydrovinylation (HV) reactions are excellent, and selectivities are in the range of 92-99% for the desired isomers. Discovery of novel, configurationally fluxional, yet sterically less demanding 2,2'-biphenol-derived phosphoramidite Ni complexes (fully characterized by X-ray) turned out to be critical for success in several HV reactions. We also report a less spectacular yet equally important role of solvents in a metal-ammonia reduction for the installation of a key benzylic chiral center. Starting with simple oxygenated styrene derivatives, we iteratively install the various exocyclic chiral centers present in typical serrulatane [e.g., a (+)- p-benzoquinone natural product, elisabethadione, nor-elisabethadione, helioporin D, a known advanced intermediate for the synthesis of colombiasin and elisapterosin] and amphilectane [e.g., A-F, G-J, and K,L pseudopterosins] derivatives. A concise table showing various synthetic approaches to these molecules is included in the Supporting Information. Our attempts to synthesize a hitherto elusive target, elisabethin A, led to a stereoselective, biomimetic route to pseudopterosin A-F aglycones.
Assuntos
Diterpenos/síntese química , Compostos de Vinila/química , Catálise , Ciclização , Diterpenos/química , Compostos Organofosforados/química , EstereoisomerismoRESUMO
1. Topical anesthesia with benzocaine or lidocaine occasionally causes methemoglobinemia, an uncommon but potentially fatal disorder where the blood has a reduced ability to transport oxygen. Previous in vitro studies using human whole blood have shown that benzocaine causes more methemoglobin (MetHb) formation than lidocaine, and that both compounds require metabolic transformation to form the MetHb producing species. In the current investigation, the active species of benzocaine forming the MetHb was investigated. 2. HPLC analysis of benzocaine samples incubated with human hepatic S9 showed the formation of a peak with the same UV spectrum and retention time as benzocaine hydroxylamine (BenzNOH). To confirm the activity of BenzNOH, MetHb production following exposure to the compound was determined in whole human blood using an Avoximeter 4000 CO-oximeter. 3. BenzNOH produced MetHb in a concentration dependent manner without the need for metabolic activation. Benzocaine in the presence of metabolic activation required a concentration of 500 µM to produce a similar degree of MetHb formation as 20 µM BenzNOH without activation. Previous work suggested that two metabolites of lidocaine may also form MetHb; N-hydroxyxylidine and 4-hydroxyxylidine. Of these two metabolites 4-hydroxyxylidine produced the most MetHb in whole blood in vitro in the absence of metabolic activation, however BenzNOH produced up to 14.2 times more MetHb than 4-hydroxyxylidine at a similar concentration. 4. These results suggest that the ability of benzocaine to form MetHb is likely to be mediated through its hydroxylamine metabolite and that this metabolite is inherently more active than the potentially MetHb-forming metabolites of lidocaine.
Assuntos
Benzocaína/metabolismo , Lidocaína/metabolismo , Metemoglobina/metabolismo , Acetaminofen/análogos & derivados , Anestésicos Locais/metabolismo , Humanos , MetemoglobinemiaRESUMO
N-sulfonated oversulfated chondroitin sulfate (NS-OSCS), recently reported as a potential threat to the heparin supply, was prepared along with its intermediate derivatives. All compounds were spiked into marketplace heparin and subjected to United States Pharmacopeia (USP) identification assays for heparin (proton nuclear magnetic resonance [(1)H NMR], chromatographic identity, % galactosamine [%GalN], anti-factor IIa potency, and anti-factor Xa/IIa ratio). The U.S. Food and Drug Administration (FDA) strong-anionic exchange high-performance liquid chromatography (SAX-HPLC) method resolved NS-OSCS from heparin and OSCS and had a limit of detection of 0.26% (w/w) NS-OSCS. The %GalN test was sensitive to the presence of NS-OSCS in heparin. Therefore, current USP heparin monograph tests (i.e., SAX-HPLC and %GalN) detect the presence of NS-OSCS in heparin.
Assuntos
Anticoagulantes/química , Sulfatos de Condroitina/análise , Contaminação de Medicamentos , Heparina/química , Indicadores e Reagentes/análise , Resinas de Troca Aniônica , Anticoagulantes/farmacologia , Sulfatos de Condroitina/química , Sulfatos de Condroitina/toxicidade , Cromatografia Líquida de Alta Pressão , Dimetilformamida/química , Contaminação de Medicamentos/prevenção & controle , Galactosamina/análise , Heparina/farmacologia , Hidrazinas/química , Indicadores e Reagentes/química , Indicadores e Reagentes/toxicidade , Limite de Detecção , Espectroscopia de Prótons por Ressonância Magnética , Controle de Qualidade , Estados Unidos , United States Food and Drug AdministrationRESUMO
Impurities are present in any drug substance or drug product. They can be process-related impurities that are not completely removed during purification or are formed due to the degradation of the drug substance over the product shelf-life. Unlike the drug substance, impurities generally do not have beneficial effects and may present a risk without associated benefit. Therefore, their amount should be minimized. 2-Bromo-3'-chloropropiophenone (BCP) is an impurity of bupropion, a second-generation antidepressant and a smoking cessation aid. The United States Pharmacopeia recommends an acceptable level for BCP that is not more than 0.1% of the bupropion. Because exposure to genotoxic impurities even at low levels is of significant concern, it is important to determine whether or not BCP is genotoxic. Therefore, in this study the Ames test and the in vitro micronucleus assay were conducted to evaluate the genotoxicity of BCP. BCP was mutagenic with S9 metabolic activation, increasing the mutant frequencies in a concentration-dependent manner, up to 22- and 145-fold induction over the controls in Salmonella strains TA100 and TA1535, respectively. BCP was also positive in the in vitro micronucleus assay, resulting in up to 3.3- and 5.1-fold increase of micronucleus frequency for treatments in the absence and presence of S9, respectively; and 9.9- and 7.4-fold increase of aneuploidies without and with S9, respectively. The addition of N-acetyl-l-cysteine, an antioxidant, reduced the genotoxicity of BCP in both assays. Further studies showed that BCP treatment resulted in induction of reactive oxygen species (ROS) in the TK6 cells. The results suggest that BCP is mutagenic, clastogenic, and aneugenic, and that these activities are mediated via generation of reactive metabolites.
Assuntos
Propiofenonas/toxicidade , Acetilcisteína/farmacologia , Linhagem Celular Tumoral , Interações Medicamentosas , Humanos , Testes de Mutagenicidade/métodosRESUMO
In response to a published concern about the potency and quality of generic vancomycin products, the United States Food and Drug Administration investigated a small sampling of the vancomycin products available in North America with regard to purity, content, and potency. To facilitate identification of impurities, a new liquid chromatography method was developed using high-resolution mass spectrometry in addition to diode array detection to characterize impurities in several commercial products. Furthermore, a microbiological assay was utilized to link the analytical profiles with an in vitro potency. All products tested met the quality specifications outlined in the United States Pharmacopeia (USP) (vancomycin hydrochloride for injection monograph) for impurities and potency (USP, Vancomycin hydrochloride for injection. United States Pharmacopeia and National Formulary, vol USP 34-NF 29, 2011).
Assuntos
Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Qualidade de Produtos para o Consumidor , Controle de Qualidade , Estados Unidos , VancomicinaRESUMO
Cationic amphiphilic drugs and aminoglycoside antibiotics can induce phospholipidosis (PLD), an abnormal accumulation of phospholipids in lysosome-derived vesicles, in preclinical studies. The incidence of PLD in patients and its clinical relevance are difficult to assess without noninvasive biomarkers. Di-docosahexaenoyl bis(monoacylglycerol)phosphate (di-22:6-BMP) is a phospholipid that is enriched in lysosomal membranes and a proposed urinary biomarker of drug-induced PLD. The specificity of di-22:6-BMP for PLD was compared to other phospholipid species that can increase in urine with nephrotoxicity. Using liquid chromatography coupled to mass spectrometry, 12 phospholipids were assayed in the urine of rats treated with drugs that induced PLD or caused renal or skeletal muscle injury. In receiver operating curve analyses, urinary di-22:6-BMP was a significantly better predictor of PLD and the least predictive of tissue injury of the phospholipids assayed. The data provide evidence supporting the use of di-22:6-BMP as a urinary biomarker of PLD in rats.
Assuntos
Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Nefropatias/induzido quimicamente , Lisofosfolipídeos/urina , Fosfolipídeos/urina , Animais , Biomarcadores/urina , Moléculas de Adesão Celular/urina , Cisplatino/efeitos adversos , Feminino , Gentamicinas/efeitos adversos , Hexestrol/efeitos adversos , Hexestrol/análogos & derivados , Nefropatias/patologia , Nefropatias/urina , Lipocalina-2 , Lipocalinas/urina , Fígado/efeitos dos fármacos , Fígado/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Linfonodos/efeitos dos fármacos , Linfonodos/patologia , Masculino , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Osteopontina/urina , Ratos , Ratos Endogâmicos F344 , Ratos Sprague-Dawley , Sinvastatina/efeitos adversos , Baço/efeitos dos fármacos , Baço/patologia , Troponina I/sangueRESUMO
In this report we highlight the significant potential of ethylene as a reagent for the introduction of a vinyl group with excellent stereoselectivity at three different stages in the synthesis of a broad class of natural products, best exemplified by syntheses of pseudopterosins. The late-stage applications of the asymmetric hydrovinylation reaction further illustrate the compatibility of the catalyst with complex functional groups. We also show that, depending on the choice of the catalyst, it is possible to either enhance or even completely reverse the inherent diastereoselectivity in the reactions of advanced chiral intermediates. This should enable the synthesis of diastereomeric analogs of several classes of medicinally relevant compounds that are not readily accessible by the existing methods, which depend on 'substrate control' for the installation of many of the chiral centers, especially in molecules of this class.
Assuntos
Alcenos/química , Antozoários/química , Diterpenos/síntese química , Etilenos/química , Glicosídeos/síntese química , Alcenos/síntese química , Animais , Etilenos/síntese química , EstereoisomerismoRESUMO
In this work we describe a 96-well microplate assay for oversulfated chondroitin sulfate A (OSCS) in heparin, based on a water-soluble cationic polythiophene polymer (3-(2-(N-(N'-methylimidazole))ethoxy)-4-methylthiophene (LPTP)) and heparinase digestion of heparin. The assay takes advantage of several unique properties of heparin, OSCS, and LPTP, including OSCS inhibition of heparinase I and II activity, the molecular weight dependence of heparin-LPTP spectral shifts, and the distinct association of heparin fragments and OSCS to LPTP. These factors combine to enable detection of the presence of 0.003% w/w spiked OSCS in 10 µg of heparin sodium active pharmaceutical ingredient (API) using a plate reader and with visual detection to 0.1% levels. The same detection limit for OSCS was observed in the presence of 10% levels of dermatan sulfate (DS) or chondroitin sulfate A (CSA) impurities. In addition, we surveyed a selection of crude heparin samples received by the agency in 2008 and 2009 to determine average and extreme DS, CSA, and galactosamine weight percent levels. In the presence of these impurities and the variable heparin content in the crude heparin samples, spiked OSCS was reliably detected to the 0.1% w/w level using a plate reader. Finally, authentically OSCS contaminated heparin sodium API and crude samples were distinguished visually by color from control samples using the LPTP/heparinase test.
Assuntos
Sulfatos de Condroitina/análise , Sulfatos de Condroitina/química , Colorimetria/métodos , Heparina/química , Microtecnologia/métodos , Sulfatos/química , Flavobacterium/enzimologia , Heparina/metabolismo , Heparina Liase/metabolismo , Polímeros/química , Polissacarídeo-Liases/metabolismo , Tiofenos/químicaRESUMO
A mixture of α-cyano-4-hydroxycinnamic acid and 1,5-diaminonaphthalene was discovered as a novel binary matrix for the qualitative analysis of 14 small-molecule (~250-550 Da) cardiovascular drugs by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and MS/MS in either positive or negative ion mode.
RESUMO
Ion mobility spectrometry was used as a rapid screening tool for the detection of acetildenafils, sildenafils and avanafil within adulterated herbal supplement matrices. Acetildenafils show a tendency for partial fragmentation during the desorption/ionization process affording two peaks in the ion mobility spectrum in addition to the intact compound. The fragmentation appears to occur α to the carbonyl group along the CN bond attaching the piperazine moiety, producing a common fragment (K0=1.0280 cm²V⻹s⻹) along with the respective piperazine fragment. The sildenafils and avanafil afford one molecular ion peak per compound.
Assuntos
Suplementos Nutricionais/análise , Contaminação de Alimentos , Inspeção de Alimentos/métodos , Inibidores da Fosfodiesterase 5/análise , Piperazinas/análise , Pirimidinas/análise , Sulfonas/análise , Vasodilatadores/análise , Carbolinas/análise , Carbolinas/química , Técnicas de Química Analítica , Imidazóis/análise , Imidazóis/química , Isomerismo , Inibidores da Fosfodiesterase 5/química , Piperazinas/química , Purinas/análise , Purinas/química , Pirimidinas/química , Citrato de Sildenafila , Sulfonas/química , Tadalafila , Espectrometria de Massas em Tandem , Triazinas/análise , Triazinas/química , Dicloridrato de Vardenafila , Vasodilatadores/químicaRESUMO
A novel analogue of sibutramine, 11-desisobutyl-11-benzylsibutramine, has been discovered. During routine ion mobility spectrometry (IMS) screening of a weight loss supplement collected at an US FDA import operation facility an unknown peak was observed. Further analysis of the supplement by liquid chromatography-mass spectrometry (LC-MS) and high resolution mass spectrometry revealed an unknown peak with a relative retention time of 1.04 with respect to sibutramine and a predicted formula of C20H24NCl. In order to elucidate the analogue's structure, it was isolated from the supplement and characterized by tandem mass spectrometry and nuclear magnetic resonance (NMR), which revealed the analogue possessed a benzyl moiety at the 11 position in place of the isobutyl group associated with sibutramine.
Assuntos
Ciclobutanos/química , Redução de Peso/efeitos dos fármacos , Cromatografia Líquida/métodos , Suplementos Nutricionais , Espectroscopia de Ressonância Magnética/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
Phospholipidosis (PLD), an abnormal accumulation of phospholipids within tissues, is observed during the preclinical testing of many pharmaceutical drugs. Diagnosis of PLD is currently based on morphologic criteria. An understanding of the clinical incidence of PLD and its possible relationship to adverse drug reactions has been hampered by the absence of noninvasive biomarkers for PLD. The uncommon phospholipid di-docosahexaenoyl bis(monoacylglycerol) phosphate (di-22:6-BMP) has been proposed as a potential urinary biomarker for PLD, but data on the utility of serum di-22:6-BMP measurements in diagnosing PLD is more limited. In this report, we compared the performance of serum and urinary di-22:6-BMP as biomarkers for PLD in rats treated with the PLD-inducing drugs amiodarone and 4,4'-diethylaminoethoxyhexestrol dihydrochloride or the hepatotoxicant acetaminophen (APAP). Serum levels of di-22:6-BMP showed a higher correlation to a generalized PLD incidence score than did levels in urine, but were unexpectedly elevated in rats with marked levels of APAP-induced liver necrosis. When samples were filtered based on serum ALT or liver histopathology thresholds, the diagnostic accuracy of serum di-22:6-BMP for PLD improved to the high level observed for urinary di-22:6-BMP without sample exclusion. These data help define the potential context-of-use of serum di-22:6-BMP as a non-clinical biomarker of PLD.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/urina , Lipidoses/sangue , Lipidoses/urina , Lisofosfolipídeos/sangue , Lisofosfolipídeos/urina , Acetaminofen/toxicidade , Amiodarona/toxicidade , Animais , Biomarcadores/sangue , Biomarcadores/urina , Doença Hepática Induzida por Substâncias e Drogas/patologia , Histocitoquímica , Masculino , Microscopia Eletrônica de Transmissão , Fosfolipídeos/sangue , Fosfolipídeos/urina , Ratos , Ratos Endogâmicos F344RESUMO
Melamine adulteration of food and pharmaceutical products is a major concern and there is a growing need to protect the public from exposure to contaminated or adulterated products. One approach to reduce this threat is to develop a portable method for on-site rapid testing. We describe a universal and selective method for the detection of melamine in a variety of solid matrices at the 100-200 µg L(-1) level by surface enhanced Raman spectroscopy (SERS) with gold nanoparticles. With minimal sample preparation and the use of a portable Raman spectrometer, this work will lead to field-based screening for melamine adulteration. Citrate coated gold nanoparticles (Au NPs) were investigated for both colorimetric and Raman-based responses. Several non-hazardous solvents were evaluated in order to develop a melamine extraction procedure safe for field applications. Au NP agglomerates formed by the addition of isopropanol (IPA) prior to sample introduction enhanced the Raman signal for melamine and eliminated matrix interference for substrate formation. The melamine Raman signal resulted in a 10(5) enhancement through the use of Au NP agglomerates. To our knowledge, we have developed the first portable SERS method using Au NPs to selectively screen for the presence of melamine adulteration in a variety of food and pharmaceutical matrices, including milk powder, infant formula, lactose, povidone, whey protein, wheat bran and wheat gluten.
Assuntos
Análise de Alimentos/métodos , Ouro/química , Nanopartículas/química , Análise Espectral Raman/métodos , Triazinas/análise , Animais , Ácido Cítrico/química , Humanos , Lactente , Fórmulas Infantis/química , Leite/química , Nanopartículas/ultraestrutura , Sensibilidade e Especificidade , Triazinas/isolamento & purificaçãoRESUMO
Ion mobility spectrometry (IMS) served as a rapid, qualitative screening tool for the analysis of adulterated weight-loss products. We have previously shown that sibutramine extracted into methanol from dietary supplements can be detected at low levels (2ng) using a portable IMS spectrometer, and have adapted a similar method for the analysis of additional weight-loss product adulterants. An FDA collaborative study helped to define the limits for fluoxetine with a limit of detection of 2ng. We also evaluated more readily available, less toxic extraction solvents and found isopropanol and water were comparable to methanol. Isopropanol was favored over water for two reasons: (1) water increases the analysis time and (2) aqueous solutions were more susceptible to pH change, which affected the detection of sibutramine. In addition to sibutamine and fluoxetine, we surveyed 11 weight-loss adulterants; bumetanide, fenfluramine, furosemide, orlistat, phenolphthalein, phentermine, phenytoin, rimonabant, sertraline and two sibutramine analogs, desmethylsibutramine and didesmethylsibutramine, using portable and benchtop ion mobility spectrometers. Out of these 13 active pharmaceutical ingredients (APIs), portable and benchtop ion mobility spectrometers were capable of screening products for 10 of these APIs. The developed procedure was applied to two weight-loss dietary supplements using both portable and benchtop instruments. One product contained didesmethylsibutramine while the other contained didesmethylsibutramine and phenolphthalein.
Assuntos
Fármacos Antiobesidade/química , Suplementos Nutricionais/análise , Ciência de Laboratório Médico/instrumentação , Ciência de Laboratório Médico/métodos , Análise Espectral/instrumentação , Análise Espectral/métodos , 2-Propanol/química , Concentração de Íons de Hidrogênio , Íons/química , Metanol/química , Soluções/química , Solventes/química , Água/química , Redução de Peso/efeitos dos fármacosRESUMO
New assays have been developed to detect the presence of economically motivated additives (EMAs) and poor manufacturing processes in heparin. Here, selected oversulfated glycosaminoglycans that are possible EMAs to heparin were synthesized and the utility of current (1)H NMR, SAX-HPLC or anticoagulation time protocols were evaluated for the detection of native impurities (chondroitin sulfate A or B (CSA or CSB), or heparan sulfate (HS)), or synthetic contaminants (oversulfated-(OS)-CSA, OS-CSB, OS-HS or OS-Heparin) spiked into heparin sodium active pharmaceutical ingredients (APIs). The range of w/w percent LOD values from the SAX-HPLC analysis for heparin spiked with CSA, CSB, HS, OS-CSA, OS-CSB, OS-HS, OS-Heparin and two partially oversulfated CSA analogs was 0.05-0.12%. The 500 MHz 1D-(1)H NMR spectra of heparin spiked with 1.0-10% CSA, CSB, OS-CSA, or OS-CSB showed unique signal pattern changes while the samples spiked with HS, OS-HS, OS-Heparin or partially sulfated CSA were more difficult to identify using NMR data. The ratio of anticoagulation time values obtained with factor Xa and factor IIa were found to remain within USP specifications in the presence of 10% amounts of these impurities or contaminants. In a separate test, using OS-CSA spiked API heparin samples, the factor Xa or factor IIa to USP standard ratio were found to fall below the USP 0.9 specification value in samples spiked at ca. weight percent of 15% or greater of OSCS. We conclude that the SAX-HPLC assay is the most sensitive and robust assay to identify and quantitate possible GAG-based EMAs in heparin.