Evaluation of Ilex guayusa and Piper marginatum Extract Cytotoxicity on Human Dental Pulp Mesenchymal Stem Cells.

BACKGROUND: Amelogenesis imperfecta is a hereditary disorder affecting dental enamel. Among its phenotypes, hypocalcified AI is characterized by mineral deficiency, leading to tissue wear and, consequently, dental sensitivity. Excessive fluoride intake (through drinking water, fluoride supplements, toothpaste, or by ingesting products such as pesticides or insecticides) can lead to a condition known as dental fluorosis, which manifests as stains and teeth discoloration affecting their structure. Our recent studies have shown that extracts from Colombian native plants, Ilex guayusa and Piper marginatum, deposit mineral ions such as phosphate and orthophosphate into the dental enamel structure; however, it is unknown whether these extracts produce toxic effects on the dental pulp. OBJECTIVE: To assess cytotoxicity effects on human dental pulp stem cells (hDPSCs) exposed to extracts isolated from I. guayusa and P. marginatum and, hence, their safety for clinical use. METHODS: Raman spectroscopy, fluorescence microscopy, and flow cytometry techniques were employed. For Raman spectroscopy, hDPSCs were seeded onto nanobiochips designed to provide surface-enhanced Raman spectroscopy (SERS effect), which enhances their Raman signal by several orders of magnitude. After eight days in culture, I. guayusa and P. marginatum extracts at different concentrations (10, 50, and 100 ppm) were added. Raman measurements were performed at 0, 12, and 24 h following extract application. Fluorescence microscopy was conducted using an OLIMPUS fv1000 microscope, a live-dead assay was performed using a kit employing a BD FACS Canto TM II flow cytometer, and data analysis was determined using a FlowJo program. RESULTS: The Raman spectroscopy results showed spectra consistent with viable cells. These findings were corroborated using fluorescence microscopy and flow cytometry techniques, confirming high cellular viability. CONCLUSIONS: The analyzed extracts exhibited low cytotoxicity, suggesting that they could be safely applied on enamel for remineralization purposes. The use of nanobiochips for SERS effect improved the cell viability assessment.

Anticariogenic Activity of Three Essential Oils from Brazilian Piperaceae.

The current trend toward using natural food additives, cosmetics, and medicines has motivated industries to substitute synthetic compounds for natural products. Essential oils (EOs) from medicinal plants are a well-known source of chemical compounds that display several interesting biological activities, including antimicrobial action. In this study, we investigated the antibacterial activity of EOs extracted from three Piperaceae species collected in the Brazilian Amazon region against a representative panel of cariogenic bacteria. The minimum inhibitory concentration (MIC) of the essential oils extracted from Peperomia pellucida (PP-EO), Piper marginatum (PM-EO), and Piper callosum (PC-EO) was determined against Streptococcus mutans, S. mitis, S. sanguinis, S. salivarius, S. sobrinus, Enterococcus faecalis, and Lactobacillus casei by using the microplate microdilution method. PM-EO, PC-EO, and PP-EO displayed antibacterial activity against all the tested cariogenic bacteria. PM-EO displayed the best inhibitory activity, with MIC values ranging from 50 to 500 µg/mL. The lowest MIC values were obtained for PM-EO against S. mitis (MIC = 75 µg/mL), Lactobacillus casei (MIC = 50 µg/mL), and S. mutans (MIC = 50 µg/mL). Gas chromatography mass spectrometry (GC-MS) analysis allowed the chemical composition of all the EOs to be identified. The main constituents of PM-EO, PC-EO, and PP-EO were 3,4-(methylenedioxy)propiophenone, α-pinene, and dillapiole, respectively. Finally, the compounds that were exclusively detected in PM-EO are highlighted. Our results suggest that PM-EO may be used in products for treating dental caries and periodontal diseases.

Leishmanicidal activity of Piper marginatum Jacq. from Santarém-PA against Leishmania amazonensis.

Leishmaniasis is an infectious disease that has high endemicity and is among the six parasitic diseases of higher occurrence in the world. The current treatments are limited due to their toxicity, treatment resistance and high cost which have increased the search for new substances of natural origin for its therapy. Based on this, an in vitro biological and chemical investigation was carried out to evaluate the potential of Piper marginatum against Leishmania amazonesis. P. marginatum leaves were collected to obtain the essential oil (EO) and the ethanolic extract (CE). The chemical profile of the CE and fractions was obtained by 1H NMR. The analysis of the EO chemical composition was performed by GC-MS. EO, CE and fractions were submitted to antileishmanial and cytotoxicity assays against macrophages. The chromatographic profiles of EO, CE and fractions showed the presence of phenolic compounds and terpenoids, having 3,4-Methylenedioxypropiophenone as a major compound. All P. marginatum samples showed low toxicity to macrophages. The CE and the methanolic, hexane and ethyl acetate fractions had low cytotoxicity when compared to Pentamidine. All tested samples inhibited growth of L. amazonensis promastigotes. The antileishmanial activity of EO, CE and fractions were evaluated in macrophages infected with L. (L.) amazonensis and treated with the concentrations 1, 10 and 100 µg/mL for 48 h. All samples were active, but EO and CE showed superior activity against amastigote forms when compared to the promastigote forms of L. amazonensis. This work describes for the first time the antileishmanial activity of the species P. marginatum and its cytotoxicity against macrophages, suggesting that it can be an alternative source of natural products in the phytotherapeutic treatment of leishmaniasis.

Folk medicine, phytochemistry and pharmacological application of Piper marginatum

ABSTRACT Piper marginatum Jacq., Piperaceae, is a widely distributed Neotropical species abundant in the Caribbean, exhibiting a characteristic winged petiole and a heart-shaped leaf, its two vegetative landmarks for rapid identification. The species has been employed by traditional indigenous cultures for its reputed medicinal properties. The plant is most frequently employed by local healers in Central America, the Antilles and South America, for alleviating gastrointestinal ailments, administered as a decoction or infusion for its tonic, diuretic and carminative effects. These beneficial properties may be attributed to the presence of various phytochemicals within P. marginatum, with most of the studies focusing on the essential oil of the plant. Monoterpenoids, sesquiterpenoids and phenylpropanoids of a varied chemical structure have been identified in the essential oil, while phenylalkanoids, aristolactams, amides and flavonoids have been purified by chromatographic techniques from the extracts. The biological and pharmacological examination of P. marginatum showed that the plant may be a valuable source of mosquitocidal, antifungal, antitumoral and hemostatic agents. Future bioguided research may yield biologically relevant molecules useful in medicine or agriculture.

Circadian variation of essential oil from Piper marginatum Jacq / Variaciones circadiana de aceite esencial de Piper marginatum Jacq

The composition of the essential oil from the leaves of Piper marginatum (Piperaceae) shows circadian variation and a higher yield during high solar incidence. The essential oils samples were investigated by GC-FID and GC-MS, which allowed identification of 29 compounds. Principal component analysis (PCA) and cluster analysis (CA) showed a significant quantitative variability in the chemical composition of the studied samples as well as a correlation between the oil profiles and the collection time. Two main groups were observed, with predominance of phenylpropanoids or sesquiterpenes. The predominant chemical compounds were phenylpropanoids, having as main representative (Z)-asarone and (E)-asarone.

La composición del aceite esencial de hojas de Piper marginatum (Piperaceae) muestra una variación circadiana y un mayor rendimiento durante la mayor incidencia solar. Las muestras de aceites esenciales se investigaron por GC-FID y GC-MS, lo que permitió la identificación de 29 compuestos. El análisis de componentes principales y el análisis de conglomerados mostraron una variación cuantitativa significativa en la composición química de las muestras estudiadas, así como una correlación entre los perfiles del aceite esencial y el tiempo de recolección. Se observaron dos grupos principales, con predominio de fenilpropanoides o sesquiterpenos. Los compuestos predominantes fueron los fenilpropanoides, (Z)-asarona y (E)-asarona.