Molecular Docking of Volatile Oily Constituents of Chinese Herbal Medicine Asari Radix et Rhizoma and CYP1A2 Enzyme / 中国实验方剂学杂志

Objective::To study the mechanisms of action of four volatile oil components (safrole, myristicin, methyleugenol and asarone) and the reactive metabolites of safrole and myristicin with CYP1A2. Method::The inhibitory effects of the volatile oil components of Asari Radix et Rhizoma on the human liver microsomal enzymes CYP1A2, CYP2D6, CYP2E1, CYP3A4 and CYP2C19 were screened by the " Cocktail" probe substrate method. The ability of the volatile oil components and intermediates in binding to CYP1A2 enzyme was studied by means of semi-flexible molecular docking. Result::The screening results showed that the components had a strong inhibitory effect on CYP1A2.Molecular docking scores were 3.048 7 kcal·mol-1 (safrole), 6.016 4 kcal·mol-1 (myristicin), 16.969 2 kcal·mol-1 (methyleugenol), 16.013 8 kcal·mol-1 (asarone), 23.923 3 kcal·mol-1 (safrole reactive metabolites) and 25.594 3 kcal·mol-1 (myristicin reactive metabolites). Conclusion::Molecular docking results indicate that safrole metabolic intermediate and myristicin metabolic intermediate have the strongest ability in binding to CYP1A2 enzyme. This study further confirms that safrole and myristicin are the mechanism-based inhibitors of CYP1A2 enzyme, which is consistent with the results of previous IC50-shift and glutathione capture experiments.

First phytochemical description of essential oils from Piper cachimboense (Piperales, Piperaceae) / Primeira descrição fitoquímica de óleos essenciais de Piper cachimboense (Piperales, Piperaceae)

Piper cachimboense is recorded only for the Amazon region of Brazil and Colombia, and the objective of this study was to report the first phytochemical assessment of the composition of the essential oils (EOs) from this species collected in the Amazon rainforest, in Novo Progresso, Pará State, Brazil. Samples of leaves were subjected to hydrodistillation in a Clevenger-type apparatus. The chemical identification was carried out by gas chromatography. The yield of oils was of 11.03 ± 5.94% for fresh leaves, and 1.07 ± 0.27% for dry leaves. The analysis showed 36 volatile compounds from fresh leaves and 49 from dried leaves. Main constituents in EOs of both fresh and dried leaves from P. cachimboense were (E)-caryophyllene, germacrene-D, -amorfene, -cadinene and apiole.

Piper cachimboense é registrada apenas para a região amazônica do Brasil e Colômbia, e o objetivo deste estudo foi relatar, pela primeira vez, a composição fitoquímica dos óleos essenciais (OEs) desta espécie coletada na floresta amazônica, em Novo Progresso/PA, Brasil. Amostras de folhas foram submetidas a hidrodestilação em aparelho tipo Clevenger. A identificação química foi realizada por cromatografia gasosa e o rendimento dos óleos foi de 11,03 ± 5,94% para as folhas frescas e de 1,07 ± 0,27% para as folhas secas. A análise mostrou 36 compostos voláteis para folhas frescas e 49 para folhas secas. Os constituintes principais dos OEs de folhas frescas e secas de P. cachimboense foram (E)-cariofileno, germacreno-D, -amorfeno, -cadineno e apiol.

Analysis on volatile components in different parts of Asarum insigne by HS-GC-MS / 中草药

Objective To perform qualitative and quantitative analysis on the volatile components in roots, rhizomes, leaves, and flowers of Asari Radix et Rhizoma derived from Asarum insigne. Methods The volatile components were analyzed by HS-GC-MS, and the relative percentage content of each component was calculated with peak area normalization method. Results There were 58 components separated from four parts of A. insigne, including 27 common components in different parts. The principal constituents was trans-β-farnesene, safrole, and asaricin. Their contents were different in four parts. Especially the contents of safrole in rhizomes, leaves, and flowers were up to 34%, 22%, and 21%; The safrole in rhizomes was over twice higher than that in roots (12%). Because safrole was extremely poisonous, the rhizomes, leaves, and flowers should be used carefully. Conclusion The volatile components in A. insigne can be detected by HS-GC-MS simply and quickly. The research can be helpful for development and quality evaluation of A. insigne.

Historical evolution and rational use of Asari Radix et Rhizoma / 中草药

Asari Radix et Rhizoma has been used for more than 2000 years with wide application in medicine. In recent years, there are many poisoning cases of Asari Radix et Rhizoma being reported in clinic, the rational application in clinic of Asarum Radix et Rhizoma has been widely concerned due to its poisonous componets of safrole and aristolochic acid. The present paper traces the historical evolution of Asari Radix et Rhizoma through accessing Web of Science and multiple databases for biomedicinal sciences, and analyzes the characteristics of its toxic reaction, and discusses the matters needing attention in its rational application from the perspective of chemical composition, so as to promote the safe and reasonable application of Asari Radix et Rhizoma.

Growth, photosynthetic pigments and production of essential oil of long-pepper under different light conditions

ABSTRACT Piper hispidinervum C. DC. is popularly known as long-pepper and it owns a commercial value due to the essential oil it produces. Long-pepper oil is rich in safrole and eugenoln components that have insecticidal, fungicidal and bactericidal activity. It has been establish that to medicinal plants light influences not only growth but also essential oil production. The growth, the content of photosynthetic pigments and the essential oil production of Piper hispidinervum at greenhouses with different light conditions was evaluated. The treatments were characterized by cultivation of plants for 180 days under different light conditions, produced by shading greenhouses with 50% and 30% of natural incident irradiance, two colored shading nets red (RN) and blue (BN) both blocking 50% of the incident radiation and one treatment at full-sun (0% of shade). The results showed that the treatments of 50% shade and RN and BN were the ones which stimulated the greater growth. Blue and red light also had the best production of photosynthetic pigments. Essential oil yielded more under full sun therefore this is the most indicated condition to produce seedlings for the chemical and pharmaceutical industry.

Metabolism and antifungal activity of safrole, dillapiole, and derivatives against botryodiplodia theobromae and colletotrichum acutatum / Metabolismo y actividad antifúngica de safrol, dilapiol, y derivados contra botryodiplodia theobromae y colletotrichum acutatum

The direct in vitro fungitoxicity and metabolism of safrole and dillapiole (isolated from Piper auritum and Piper holtonii, respectively) by Botryodiplodia theobromae and Colletotrichum acutatum were investigated. Higher values of mycelial growth inhibition for both fungi were obtained for dillapiole, as compared with safrole. B. theobromae was able to metabolize both compounds to their respective vicinal diols, reaching 65 percent relative abundance during the biotransformation of dillapiole; while C. acutatum only transformed safrole to various metabolites with relative abundances under 5 percent. According to the low antifungal activity of the major metabolic products (< 5 percent for vicinal diols), a detoxification process was implied. Studies on the influence of some substituents in the aromatic ring of safrole and dillapiole on the antifungal activity against B. theobromae were also carried out. As result, the safrole nitrated derivative, 6-nitrosafrole, showed a fungitoxicity level similar to that displayed by the commercial fungicide Carbendazim® under the conditions used. In light of this, safrole and dillapiole could be suggested as feasible structural templates for developing new antifungal agents.

Se investigó la fungitoxicidad directa in vitro y el metabolismo de safrol y dilapiol (obtenidos desde Piper auritum and Piper holtonii, respectivamente) por Botryodiplodia theobromae y Colletotrichum acutatum. Los valores mayores de inhibición del crecimiento micelial de ambos hongos se obtuvieron para dilapiol, en comparación con safrol. B. theobromae metabolizó ambos compuestos a sus respectivos dioles vecinales, alcanzando abundancias relativas del 65 por ciento durante la biotransformación del dilapiol; mientras que C. acutatum solo transformó safrol en varios metabolitos con abundancias relativas menores al 5 por ciento. De acuerdo con la baja actividad antifúngica de los productos metabólicos mayoritarios (< 5 por ciento para los dioles vecinales), se sugiere un proceso de desintoxicación. Adicionalmente, se evaluó la influencia de algunos sustituyentes en el anillo aromático de safrol y dilapiol sobre la actividad antifúngica contra B. theobromae. Como resultado, el derivado nitrado del safrol, el 6–nitro safrol, presentó un nivel de fungitoxicidad similar al exhibido por el fungicida comercial Carbendazim® bajo las condiciones usadas. A la luz de lo anterior, safrol y dilapiol podrían ser sugeridos como plantillas estructurales adecuadas para el desarrollo de nuevos agentes antifúngicos.

Research on reducing safrole and aristolochic acid A in Asari Radix et Rhizoma based on different processing techniques / 中草药

Objective: To study the effect of processing techniques on reducing the contents of safrole and aristolochic acid A in Asari Radix et Rhizoma. Methods: The contents of safrole and aristolochic acid A between raw herb and processed products were determined by HPLC, and the differences in the contents of toxic components between raw herb and processed products were used to evaluate the detoxification efficiency. Results: The linear ranges of safrole and aristolochic acid A were 169.2-846.0 μg (r = 0.9996) and 11.6-58.0 ng (r = 0.9996), respectively. And the average recovery rates were 99.83% (RSD = 1.67%) and 101.43% (RSD = 1.25%). The removal rate of safrole in order was as follows: salt system > fried coke > rice water system > alkali > liquorice > vinegar > ginger > wine > alkali-vinegar > honey and the removal rate of aristolochic acid A was in ordor as: fried coke > alkali-vinegar > salt system > alkali > vinegar > rice water system > liquorice > wine > honey > ginger. The removal rate of aristolochic acid A in Asari Radix et Rhizoma by fried coke was over 60%. Conclusion: The contents of safrole and aristolochic acid A in Asari Radix et Rhizoma could be decreased to different extent by processing techniques. Among the processing methods, processing by fried coke is the best method.

Estudo da adaptação da espécie Piper hispidinervum C. DC. (pimenta longa) à região do Vale do Itajaí - SC, através da composição química do óleo essencial obtido por hidrodestilação por micro-ondas e convencional / Adaptation studies of Piper hispidinervum C. DC. (long pepper) species in Itajaí Valley - SC, by the chemical composition of essential oil obtained by microwave and traditional hydrodistillation

Neste trabalho, a espécie Piper hispidinervum (pimenta longa), endêmica do estado do Acre, foi cultivada na região do Vale do Itajaí - SC, e sua adaptação foi avaliada em relação à composição química do óleo essencial obtido pelos processos de hidrodestilação convencional e por micro-ondas. O safrol foi identificado como o constituinte majoritário do óleo essencial desta espécie, o qual foi empregado como parâmetro de avaliação da capacidade de adaptação desta planta à região sul do Brasil, já que a proposta abrange avaliar a utilização desta espécie como fonte alternativa de safrol, em substituição a Canela Sassafrás (Ocotea odorifera), espécie muito explorada até a década de 90 nesta região. As amostras da planta foram obtidas de diferentes regiões do Acre e foram cultivadas na estação experimental da EPAGRI - Itajaí-SC. O óleo essencial das folhas forneceu um teor médio de safrol entre 76,6% e 89,9%. A análise por CG-DIC e CG-EM do óleo com maior concentração de safrol, apresentou os seguintes constituintes: safrol (89,93%), α-terpineno (0,35%), (E)-ß-ocimeno (0,54%), terpinoleno (3,10%), valenceno (0,21%), (Z)-ß-bisaboleno (1,70%) e guaiol (0,29%).

In this work a Piper hispidinervum specie (long pepper), endemic from the Acre state, was grown in Itajaí Valley in Santa Catarina State, and its adaptation was evaluated in relation to chemical composition of essential oil obtained by traditional hydrodistillation and microwave hydrodistillation. The safrole was identified as the major constituent of the essential oil of this specie, which was used as a measure of evaluation of the adaptation capacity of this plant in the southern Brazil, as the proposal includes the evaluation of the usage of this specie as an alternative source of safrole, replacing Sassafras Cinnamon (Ocotea odorifera), widely explored specie in this region until the 90's decade. The plant samples were obtained from different regions of the Acre state and were cultivated at the experimental station of EPAGRI Itajaí - SC. The essential oil of the leaves provided an average content of safrole between 76.6% and 89.9%. The analysis by GC-FID and GC-MS of the oil with higher safrole content, presented the following relative constitution: safrole (89.93%), α-terpinene (0.35%), (E)-ß-ocimene (0.54%), terpinolene (3.10%), valencene (0.21%), (Z)-ß-bisabolene (1.70%) and guaiol (0.29%).

Isomerização do óleo essencial de pimenta-longa (Piper hispidinervium C. DC) para a obtenção de isosafrol / Isomerization of essential oil of (Piper hispidinervium) C. DC to obtain isosafrole

As piperáceas brasileiras apresentam propriedades para serem utilizadas, entre outras aplicações, como inseticidas e antifúngicos. Dentre as piperáceas, pode-se citar a Piper hispidinervum C. DC. De suas folhas e dos talos finos extrai-se um óleo essencial rico em safrol, que é utilizado pela indústria como matéria-prima na manufatura, por exemplo, do piperonal. A sequência natural do processo para a obtenção do piperonal é o de submeter o safrol obtido da concentração do óleo essencial à isomerização para seu correspondente isômero estável chamado isosafrol. Apresenta-se neste artigo, por sua vez, a síntese do isosafrol diretamente do óleo essencial de pimenta-longa (Piper hispidinervium C. DC). Este óleo essencial apresenta o safrol como constituinte majoritário, possibilitando a sua isomerização para a produção de isosafrol, que é empregado nas indústrias farmacêuticas e de fragrâncias. O objetivo deste trabalho é o de apresentar a obtenção do isosafrol sem a necessidade da etapa de separação do safrol do óleo essencial de pimenta-longa. Para tanto, foram realizados ensaios de isomerização do óleo essencial, obtendo-se uma solução contendo 79,4 % da mistura cis, trans-isosafrol. Ressalte-se que o óleo essencial continha 86,4 % de safrol, resultando em alto rendimento na sua conversão a cis, trans-isosafrol de 97,1 %.

The Brazilian peppers present proprieties that can be used in fragrance and pharmaceutical industries, for example. One these pepper is long pepper (Piper hispidinervium C. DC.), who its essential oil is rich in safrole. Safrole is an important compound with application in several industries, such as pharmaceutics, cosmetics. After its chemical conversion, for example, it is possible to obtain heliotropin, a fixative agent for fragrances. The sequence to obtain the heliotropin is by safrole isomerization, that, initially, produce isosafrole. In this work, however, presents the synthesis of isosafrole directly from essential oil of Piper hispidinervium C. DC. In this case, the isomerization of essential oil was done getting 79,4 % of cis, trans-isosafrole mixture. The essential oil presented 86,4 % of safrole, resulting in 97,1 % yield to cis, trans-isosafrole.

Atividade inseticida do óleo essencial de pimenta longa (Piper hispidinervum C. DC.) sobre lagarta-do-cartucho do milho Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera: Noctuidae) / Insecticidal Activity of Long-pepper essential oil (Piper hispidinervum C. DC.) on fall armyworm Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera: Noctuidae)

Os objetivos desta pesquisa foram a obtenção e caracterização do óleo essencial de folhas de pimenta longa Piper hispidinervum, e avaliação de seu efeito no comportamento e/ou mortalidade da lagarta-do-cartucho do milho Spodoptera frugiperda. O óleo essencial foi obtido pela técnica "arraste a vapor d'água", utilizando-se de um aparelho de Clevenger modificado, e posteriormente submetido, à análise por CG-EM e CG. Foram realizados testes de ingestão e contato tópico em lagartas de 1º e 3º ínstar. Os resultados constataram que o óleo essencial de pimenta-longa possui atividade inseticida sobre S. frugiperda, causando redução alimentar e mortalidade, sendo o safrol (82 por cento) seu constituinte majoritário. Verificou-se mortalidade no teste de ingestão em lagartas de 1º ínstar com CL50 = 16,2 mg/mL e para lagartas de 3º ínstar a CL50 = 9,4 mg/mL com redução alimentar CD50 = 0,72 mg/mL; e de toxicidade aguda no teste de contato tópico com DL50 = 277,91 μg/lagarta, após o intervalo de tempo de 96 horas, sendo também observados sintomas de neurotoxicidade, como o efeito knock-down.

The objectives of this research were to evaluate the chemical composition and bioactivity of the essential oil from the long-pepper leaves (Piper hispidinervum) and its effect on the comportment and/or mortality of the fall armyworm (Spodoptera frugiperda). The essential oil was obtained by steam stream distillation, using the modified Clevenger apparatus, the chemical composition was analyzed by GC-MS and GC. Tests of ingestion and topical application in fall armyworm of 1 st and 3 rd ínstar were made. The results showed that the long-pepper essential oil has insecticide activity on S. frugiperda, causing mortality and food reduction, and safrole (82 percent) as its majority constituent. Mortality was found in the ingestion test in larvae of 1 st ínstar LC50 = 16.2 mg/mL for larvae of 3 rd ínstar LC50 = 9.4 mg/mL, with reduced food CD50 = 0.72 mg/mL; and acute toxicity test of the contact topic with LD50 = 277.91 μg/larvae, after an interval of 96 hours, neurotoxicity symptoms were also observed, as knock-down effect.

Efeito do óleo essencial de pimenta longa (Piper hispidinervum C. DC) e do emulsificante Tween® 80 sobre o crescimento micelial de Alternaria alternata (Fungi: Hyphomycetes) / The effect of Long-pepper essential oil (Piper hispidinervum C. DC.) and of Tween®; 80 emulsifier on the mycelial growth of Alternaria alternate (Fungi: Hyphomycetes)

Este trabalho objetivou avaliar o efeito do óleo essencial de folhas de pimenta longa (Piper hispidinervum C. DC), sobre o crescimento micelial de Alternaria alternata e a análise da influência da concentração do emulsificante Tween® 80 no controle deste fitopatógeno. O óleo essencial foi obtido pela técnica "arraste a vapor d'água", utilizando-se aparelho de Clevenger modificado, e posteriormente submetido, à análise por cromatografia em fase gasosa acoplada a espectrômetro de massas CG-EM a CG. Para os ensaios biológicos, o método foi o bioanalítico in vitro observando-se o crescimento ou inibição do micélio de A. alternata no meio de cultura BDA na presença de diferentes concentrações do óleo essencial (0, 100, 250, 500 e 1000 mgL-1 ) sob diferentes concentrações de Tween® 80. Adotou-se esquema fatorial com quatro repetições, em delineamento inteiramente casualizado (DIC). Observou-se que o óleo essencial de pimenta longa apresentou inibição sobre o crescimento micelial do fungo A. alternataem todas as concentrações analisadas, sendo que na concentração de 1000 mgL-1 esta inibição foi de 100 por cento , e a porcentagem de emulsificante (Tween® 80), influenciou na atividade fungitóxica das concentrações de 250 mgL -1 e 500 mgL -1 do óleo essencial.

The object of this work was to evaluate the effectof essential oils from Long-pepper leaves Piper hispidinervum on the mycelial growth of Alternaria alternata and to analyze the influence of emulsifier Tween® 80 concentration in the control of this phytopathogen. The essential oil was obtained by steam stream distillation, using a modified Clevenger apparatus, and the chemical composition was analyzed by gas chromatography coupled to mass spectroscopy GC-MS and GC. For biological tests, the bioanalytical in vitro method was used, observing the growth or inhibition of such phytopathogen in BDA culture media, in the presence of different essential oil concentrations (0, 100, 250, 500, e 1000 mgL -1) under different Tween® 80 concentrations. Factorial scheme with four repetitions, in entirely randomized outline was adopted. The essential oil of Long-pepper inhibited the micellial growth of the A. alternata fungus at all of the concentrations tested. A 100 percent inhibition at the 1000 mgL -1 concentration was observed, and the percentage of emulsifier (Tween® 80) influenced the fungitoxic activity at the 250 mgL-1 and 500 mgL -1 concentrations of the essential oil.

Caracterização e avaliação produtiva de uma população nativa de pimenta longa (Piper hispidinervum C.DC) no Seringal Cachoeira, AC, Brasil

The search for economic alternatives to traditional extractivism in Amazonia is important and the exploration of natural populations of plants of the Piperaceae family, which is abundant in Acre, can be a profitable activity. The objective of this study was to determine some demographic, ecological and yield characteristics of the species long pepper (Piper hispidinervum C.DC) and to evaluate the possibility of its management for the production of essential oil, with high safrole content. The study was done in the Chico Mendes Extractive Project, Xapuri, Acre, in a ten year old fallow, where long pepper was dominant. The population was evaluated through a 12% intensity random sample, with sampling plots of 10 10 m. A mean density of 15 individuals/100 m2 was verified, varying from zero to 71 individuals. This population presented an average safrole content of 92% in the essential oil, with CV=2.38%. The mean yield of oil in the dry biomass was 3.5%, with CV=28.57%. Oil yield was estimated at 12,11 kg.ha-1, with a single harvest/year, which represents 15,1% of the yield obtained in cultivation. The viability of the exploitation of these populations depends on the development of techniques to increase plant density. Furthermore, in the choice of populations for management, size, density and age of the population should be considered.

A busca de alternativas econômicas ao extrativismo tradicional na Amazônia é importante e a exploração de populações naturais de plantas da família Piperaceae, abundantes no Acre, pode se constituir em uma atividade rentável. O objetivo deste trabalho foi determinar algumas características demográficas, ecológicas e produtivas da espécie pimenta longa (Piper hispidinervum C.DC) e avaliar a possibilidade de seu manejo para a produção de óleo essencial com alto teor de safrol. O estudo foi desenvolvido no Projeto de Assentamento Extrativista Chico Mendes, em Xapuri, AC, em uma área de capoeira com aproximadamente dez anos, onde a pimenta longa era dominante. A população foi avaliada por meio de amostragem aleatória simples, com intensidade de 12%, tendo como unidades amostrais parcelas de 10 10 m. Constatou-se uma densidade média de 15 indivíduos/100 m2, variando de zero a 71 indivíduos. Esta população apresentou uma média de 92% de teor de safrol no óleo essencial, com CV de 2,38%. O rendimento médio de óleo da biomassa seca, foi de 3,5%, com CV de 28,57%. A produção de óleo foi estimada em 12,11 kg/ha, com um corte por ano, que representa apenas 15,1% da produtividade obtida em áreas de cultivo. Portanto, a viabilidade da exploração dessas populações depende do desenvolvimento de técnicas que permitam aumentar a densidade de indivíduos. Ademais, na escolha de populações para o manejo, deve-se considerar características como: tamanho, densidade e idade da população.

Determination of Safrole and 6-methyl Coumarin in Cosmetics Using GC and GC-MS / 环境与健康杂志

Objective To develop a method for separation and determination of safrole and 6-methyl coumarin in cosmetics by GC-MS. Methods The cosmetics were extracted with methanol and then dehydrated,and filtrated through the membrane of 0.45 ?m. The filtration liquid was injected into GC and GC-MS for detection and quantitative analysis with external standard method. Results The recovery rates of safrole and 6-methyl coumarin were 101.3%-106.0% and relative standard deviations of safrole and 6-methyl coumarin were 0.9%-1.4% and 0.9%-2.1% respectively. The detection limits were 5 and 10 ?g/g. Conclusion This method is accurate and sensitive for detection of safrole and 6-methyl coumarin in cosmetic.

Quaternary ammonium salt derivatives of allylphenols with peripheral analgesic effect

Ammonium salt derivatives of natural allylphenols were synthesized with the purpose of obtaining potential peripheral analgesics. These drugs, by virtue of their physicochemical properties, would not be able to cross the blood brain barrier. Their inability to enter into the central nervous system (CNS) should prevent several adverse effects observed with classical opiate analgesics (Ferreira et al., 1984). Eugenol (1) O-methyleugenol (5) and safrole (9) were submitted to nitration, reduction and permethylation, leading to the ammonium salts 4, 8 and 12. Another strategy applied to eugenol (1), consisting in its conversion to a glycidic ether (13), opening the epoxide ring with secondary amines and methylation, led to the ammonium salts 16 and 17. All these ammonium salts showed significant peripheral analgesic action, in modified version of the Randall-Sellito test (Ferreira et al. 1978), at non-lethal doses. The ammonium salt 8 showed an activity comparable to that of methylnalorphinium, the prototype of an ideal peripheral analgesic (Ferreira et al., 1984).