Metabolic products of European-type propolis. Synthesis and analysis of glucuronides and sulfates.

ETHNOPHARMACOLOGICAL RELEVANCE: Propolis is a bee-derived product used since antiquity for its general health-giving properties and is especially noted for its anti-bacterial activity. In more recent times, propolis has been employed against more specific targets such as antiproliferative effects vs cancer cells, wound healing and type-2 diabetes. AIM OF THE STUDY: European (poplar)-type propolis from New Zealand contains a number of hydroxy cinnamic acid esters and a set of aglycone flavonoid compounds, mainly chrysin, galangin, pinocembrin and pinobanksin. Propolis is usually taken orally and propolis metabolites quickly appear in the plasma of the ingested. In this work we aimed to identify the major flavonoid plasma metabolites by direct analysis of the plasma. MATERIALS AND METHODS: After consumption of a large dose of propolis in a single sitting, blood samples were taken and analysed using LCMS/MS. The major flavonoid metabolites identified were also synthesised using chemical (sulfates) or enzymatic methods (glucuronides). RESULTS: Both the sulfate and glucuronide conjugates of the four major propolis flavonoids are readily detected in human plasma after propolis ingestion. Preparation of the sulfates and glucuronides of the four major flavonoids allowed the relative proportions of the various metabolites to be determined. Although the sulfates are seen as large peaks in the LCMS/MS, the glucuronides are the dominant conjugate species. CONCLUSIONS: This study shows most of the flavonoids in the plasma are present as 7-O-glucuronides with only galangin showing some di-glucuronidation (3,7-O-diglucuronide). No evidence was found for hydroxy cinnamic acid type metabolites in the plasma samples.

Optimization and evaluation of propolis liposomes as a promising therapeutic approach for COVID-19.

The present work aimed to develop an optimized liposomal formulation for enhancing the anti-viral activity of propolis against COVID-19. Docking studies were performed for certain components of Egyptian Propolis using Avigan, Hydroxychloroquine and Remdesivir as standard antivirals against both COVID-19 3CL-protease and S1 spike protein. Response surface methodology and modified injection method were implemented to maximize the entrapment efficiency and release of the liposomal formulation. The optimized formulation parameters were as follow: LMC of 60 mM, CH% of 20% and DL of 5 mg/ml. At those values the E.E% and released % were 70.112% and 81.801%, respectively with nanosized particles (117 ± 11 nm). Docking studies revealed that Rutin and Caffeic acid phenethyl ester showed the highest affinity to both targets. Results showed a significant inhibitory effect of the optimized liposomal formula of Propolis against COVID-3CL protease (IC50 = 1.183 ± 0.06) compared with the Egyptian propolis extract (IC50 = 2.452 ± 0.11), P < 0.001. Interestingly, the inhibition of viral replication of COVID-19 determined by RT_PCR has been significantly enhanced via encapsulation of propolis extract within the liposomal formulation (P < 0.0001) and was comparable to the viral inhibitory effect of the potent antiviral (remdesivir). These findings identified the potential of propolis liposomes as a promising treatment approach against COVID-19.

Propolis-based niosomes as oromuco-adhesive films: A randomized clinical trial of a therapeutic drug delivery platform for the treatment of oral recurrent aphthous ulcers.

Oromuco-adhesive films for buccal delivery of Propolis extract (PPE) entrapped in niosomes, were prepared to treat oral recurrent aphthous ulcer (RAU). PPE was investigated for antimicrobial compounds. Niosomes composed of span60 and cholesterol were evaluated for particles size, polydispersity index (PDI), zeta-potential, entrapment efficiency and in vitro release. The formed oromuco-adhesive films containing niosomal PPE were evaluated for swelling, mucoadhesion and elasticity. 24 patients suffering from RAU were divided equally into medicated and placebo groups and participated in this study to examine the onset of ulcer size reduction, complete healing and pain relief. Ultra-performance liquid chromatography-high resolution mass spectrometry revealed the presence of pinocembrin, pinobanksin, chrysin and galangin as antimicrobial flavonoids with total content of 158.7 ± 0.15 µg quercetin equivalents and phenolic content of 180.8 ± 0.11 µg gallic acid equivalents/mg. Multilamellar niosomes of 176-333 nm displayed entrapment efficiency of 91 ± 0.48%, PDI of 0.676 and zeta potential of -4.99. In vitro release after 8 h from niosomal dispersion and films were 64.05% and 29.09 ± 0.13% respectively. Clinical results revealed duration of film adherence from 2-4 h in the two groups. The onset of ulcer size reduction in medicated group was attained within second and third day, complete healing was achieved within first 10 days of treatment and pain relief lasted for more than 4-5 h, in contrast to the placebo group. This oromuco-adhesive films which offer controlled and targeting drug delivery can be proposed as a new therapeutic strategy in the treatment of oral recurrent aphthous ulcer.

Ensayo clínico aleatorizado sobre la efectividad de tratamientos alternativos en la estomatitis aftosa recurrente / Randomized clinical trial of the effectiveness of complementary therapies for recurrent aphthous stomatitis

Fundamento y objetivo: A pesar de la elevada prevalencia de la estomatitis aftosa recurrente (EAR), su etiología no está del todo aclarada y no existe un tratamiento totalmente curativo. El objetivo de este trabajo fue evaluar la eficacia clínica y la seguridad de 4 tratamientos (nitrato de plata, própolis, ruibarbo y nogal) de la EAR. Pacientes y método: Se realizó un ensayo clínico aleatorizado con 125 pacientes con aftas menores, con 25 pacientes por grupo: cauterización con nitrato de plata, própolis, extracto de ruibarbo, extracto de corteza de nogal y placebo. Resultados y conclusiones: Ningún paciente refirió efectos adversos relacionados con el tratamiento. Hay diferencias significativas (p<0,001) globales en el tiempo hasta la desaparición de los síntomas. El más rápido fue el nitrato de plata (1,16 días), después los 3 tratamientos alternativos (1,60 días con própolis, 1,84 con ruibarbo y 2,00 con nogal, sin diferencias entre ellos), y por último el placebo (4,64 días). En cuanto al tiempo medio de curación de las lesiones, fue estadísticamente mayor (8,96 días) para el placebo que para los 4 tratamientos: nitrato de plata (7,32 días), própolis (6,80), ruibarbo (7,72) y nogal (8,00) (AU)

Background and objective: Despite the high prevalence of recurrent aphthous stomatitis (RAS), its etiology is not yet completely clear and there is no completely remedial treatment available at present. The objective of this study was to evaluate the clinical efficacy and safety of 4 treatments (silver nitrate, propolis, rhubarb and walnut) for RAS. Patients and method: A randomized clinical trial was conducted with 125 patients with minor aphthae, including 25 patients per group: cauterization with silver nitrate, propolis, rhubarb extract, walnut extract and placebo. Results and conclusions: No patient reported adverse effects related to the treatment received. There were significant (P<.001) differences in the time elapsed until symptom resolution. The fastest treatment was silver nitrate (1.16 days), followed by the 3 alternative treatments (1.60 days with propolis, 1.84 with rhubarb and 2.00 with walnut; with no differences between them), and finally the placebo (4.64 days). The mean healing time of the lesions was statistically higher (8.96 days) for the placebo than for the 4 treatments: silver nitrate (7.32 days), propolis (6.80), rhubarb (7.72) and walnut (8.00) (AU)

Propolis organogel as a novel topical delivery system for treating wounds.

CONTEXT: Propolis has traditionally been used in curing infections and healing wounds and burns. OBJECTIVE: The aim of this study is to formulate pluronic lecithin organogel of propolis to improve its availability and antimicrobial activity. MATERIALS AND METHODS: Different organogels were prepared by using soybean lecithin, isopropyl palmitate, pluronic F127 and water. The effect of quantity of lecithin and pluronic F127 and percentage of oil phase was investigated. The organogels were evaluated for appearance, texture, pH, drug content and viscosity. In vitro release studies were carried out using cellophane membrane. Drug permeation through abdominal rat skin from organogels that showed high % drug release was compared to that from propolis suspension in distilled water. Finally, the antimicrobial activity of the selected propolis formulation against different bacterial isolates was compared with that of propolis suspension in water. RESULTS AND DISCUSSION: Results showed that all organogel formulations except the formula containing 10% pluronic F127, showed acceptable physical properties. Drug content of organogel formulations was in the range of 97.5-100.2%. The pH of the formulations was in the range of 5.5-6.3 that suits the skin pH, indicating skin compatibility. The viscosity was in the range of 5366-8984 cp. A significant decrease in drug release from formulations was observed with increase in concentration of lecithin and pluronic F127. Decreasing oil phase percentage to 20% w/w led to a decrease in drug release from the formulation. CONCLUSION: The formula containing 3% lecithin and 20% pluronic F127 exhibited superior skin permeation and antimicrobial activity over propolis suspension in water.

Propolis attenuates cobalt induced-nephrotoxicity in adult rats and their progeny.

The aim of this study was to evaluate the biochemical changes in cobalt-exposed rats and to investigate the potential role of Tunisian propolis against the cobalt-induced renal damages. Twenty-four pregnant Wistar rats were divided into four groups and were treated as follows: group 1 (control) received distilled water; group 2 received 350 ppm of CoCl(2) in drinking water; group 3 received 350 ppm CoCl(2) in drinking water and a propolis-supplemented diet (1 g/100 g of diet); group 4 received a propolis-supplemented diet (1 g/100 g of diet) without cobalt. In the cobalt group, a significant decrease in body, absolute and relative weights was noted when compared to controls. The administration of cobalt to pregnant rats from the 14th day of pregnancy until day 14 after delivery resulted in an increased level of renal malondialdehyde, a decreased renal content of glutathione and antioxidant enzyme activities such as superoxide dismutase, catalase and glutathione peroxidase in lactating rats and their pups. A statistically significant increase in plasma urea and creatinine serum levels was seen in treated female rats and their pups. Histopathologically, the cobalt-administration induced degenerative changes in the kidney of lactating rats and their pups. When compared with cobalt-treated rats, those receiving the propolis supplementation (along with cobalt-treatment) had lower malondialdehyde levels, higher antioxidant activities and the cobalt-related histopathological changes in the kidneys were at lower severity. Our results suggested that the propolis might be a potential candidate agent against cobalt-induced nephrotoxicity in adult and juvenile rats when administered to female rats during the late pregnancy and the early postnatal period.

Antibacterial activity of propolis-based toothpastes for endodontic treatment / Atividade antibacteriana de pastas dentífricas com base de própolis para tratamento endodôntico

This study evaluated the antibacterial activity of propolis-based toothpastes used as intracanal medication in endodontic treatment. The propolis-based toothpastes were prepared using an extract established in previous studies (identified as A70D and D70D). Calcium hydroxide paste was used as a control. The bacteria employed were Streptococcus mutans (ATCC 25175), Staphylococcus aureus (ATCC 6538), Staphylococcus aureus (ATCC 25923), Kocuria rhizophila (ATCC 9341), Escherichia coli (ATCC 10538), Pseudomonas aeruginosa (ATCC 27853), Enterococcus hirae (ATCC 10541), Streptococcus mutans (ATCC 25175). Five field strains isolated from saliva were used: Staphylococcus spp. (23.1 - coagulase positive), Staphylococcus spp. (23.5 - coagulase negative), Staphylococcus spp. (26.1 - coagulase positive), Staphylococcus spp. (26.5 - coagulase negative) and Staphylococcus epidermidis (6epi). The diffusion-well method on double-layer agar was used in a culture medium of Tryptic Soy Agar. The plates were kept at room temperature for two hours to allow the diffusion of pastes in the culture medium, and then incubated at 35º C for twenty-four hours in aerobiosis and in microaerophilia (S. mutans). After this period, the total diameter of the inhibition halo was measured. The results were analyzed by ANOVA analysis of variance, followed by the Tukey test at p<0.05. The propolis-based toothpastes presented antibacterial activity against 83.3 percent of the analyzed bacteria. For 66.7 percent of these bacteria, the propolis-based toothpastes exhibited greater antibacterial activity than calcium hydroxide. The present results allow us to conclude that the experimental pastes A70D and D70D showed good activity against aerobic bacteria, proving more effective than calcium hydroxide.

O objetivo deste estudo foi avaliar a atividade antibacteriana de formas farmacêuticas a base de própolis para uso no tratamento endodôntico como medicação intracanal. As formulações de própolis, em forma de pastas, foram preparadas a partir de um extrato pré-estabelecido em estudos anteriores e identificadas como A70D e D70D. Como controle, foi utilizado pasta de hidróxido de cálcio. As bactérias utilizadas foram: Streptococcus mutans (ATCC 25175), Staphylococcus aureus (ATCC 6538), Staphylococcus aureus (ATCC 25923), Kocuria rhizophila (ATCC 9341), Escherichia coli (ATCC 10538), Pseudomonas aeruginosa (ATCC 27853), Enterococcus hirae (ATCC 10541), Streptococcus mutans (ATCC 25175) e 5 cepas de campo isoladas da saliva: Staphylococcus spp. (23.1 - coagulase positiva), Staphylococcus spp. (23.5 - coagulase negativa), Staphylococcus spp. (26.1 - coagulase positiva), Staphylococcus spp. (26.5 - coagulase negativa) e Staphylococcus epidermidis (6epi). Foi utilizado o método poço difusão em camada dupla de ágar, em meio de cultura Tryptic Soy Agar. As placas foram mantidas à temperatura ambiente por 2 h para permitir a difusão das pastas no meio de cultura, e então incubadas a 35 ºC por 24 h em aerobiose e em microaerofilia (S. mutans). Após este período, foi medido o diâmetro total do halo de inibição. Os resultados foram submetidos ao teste de análise de variância ANOVA seguido do teste de Tukey com p<0,05. As pastas a base de própolis apresentaram atividade antibacteriana contra 83,3 por cento das bactérias analisadas. Para 66,7 por cento das bactérias, as pastas de própolis apresentaram maior atividade antibacteriana que o hidróxido de cálcio, e este foi mais efetivo apenas para Streptococcus mutans (ATCC 25175), Staphylococcus aureus (ATCC 25923) e Pseudomonas aeruginosa (ATCC 27853). De acordo com a metodologia utilizada, pode-se concluir que as pastas experimentais A70D e D70D apresentam boa atividade contra bactérias aeróbias...

Development of topical functionalized formulations added with propolis extract: stability, cutaneous absorption and in vivo studies.

Propolis, which is a natural product widely consumed in the folk medicine, is a serious candidate to be applied topically due to its outstanding antioxidant properties. So, the purpose of this study was to develop stable topical formulations added with propolis extract in an attempt to prevent and/or treat the diseases occurring in skin caused by UV radiation. The antioxidant activity using a chemiluminescent method was used to evaluate the functional stability and the permeation/retention in skin of these formulations. In the long-term stability study, the formulations were stored at 25+/-2 degrees C/AH and at 40+/-2 degrees C/70% RH for 360 days. It was found in this study, that the formulations prepared with Polawax showed functional and physical stability in the period of study. In addition, this formulation presented good results in the percutaneous study, allowing the antioxidant compounds present in the propolis extract to reach lower layers in pig ear skin and in the whole hairless mice skin (retention=0.12 and 0.13 microL of propolis/g of skin, respectively). In the in vivo study, it was also suggested that this formulation may be effective in protecting skin from UVB photodamage, nevertheless other assays need to be done in order to have a complete understanding of the protective effect of formulations added with propolis extract.

Atividade microbiana de óleos essenciais e extratos de própolis sobre bactérias cariogênicas / Antimicrobial activity of essential oils and propolis against oral pathogens

A própolis é uma resina produzida por abelhas a partir de diferentes plantas. Os óleos essenciais são produtosdo metabolismo secundário das plantas e usados durante anos na medicina popular como antimicrobianos. Oobjetivo deste trabalho foi avaliar o perfil de susceptibilidade dos extratos etanólicos de própolis oriundos dos Estados do Paraná, Minas Gerais e São Paulo e de óleos essenciais frente às bactérias cariogênicas Streptococcus mutans e Lactobacillus casei usando o método de difusão em ágar. Os resultados obtidos indicam a própolis do estado de Minas Gerais como a mais ativa de todas, o que leva a crer que a diferença observada na quantidade de própolis produzida e a diversidade dos metabólitos secundários, no produto, podem estar associados com a região de produção e com a estação climática da coleta. Os óleos essenciais mostraram atividades significantes para os microrganismos testados, alguns deles apresentando zona de inibição maior que o do controle positivo.

Propolis is a natural composite balsam, produced by honey bees (Apis melifera) from the gum of various plants. Essential oils and their components are products of the secondary metabolism of plants, used for many ears in folk medicine, principally, as antimicrobial. The aim of the present work was to analyze the differences in the antimicrobial activity of propolis obtained from three different regions of Brazil (Minas Gerais, Parana and São Paulo States) and of essential oils, using agar diffusion methods to test them against cariogenic microorganisms Streptococcus mutans and Lactobacillus casei. The results showed that the propolis from Minas Gerais State was more active than that from the others, and implied that the existing differences in the amount of propolis produced and diversity of secondary metabolites in the product may be associated with the region of production and the climatic season of collection. The essential oils displayed significant inhibitory activity against the microorganisms tested, showing larger inhibition zones than the positive control.

Effect of Brazilian propolis on scratching behavior induced by compound 48/80 and histamine in mice.

We studied the effect of Brazilian propolis on scratching behavior induced by compound 48/80 and histamine in ICR mice. Propolis granular A.P.C dose-related inhibited scratching behavior induced by compound 48/80 and significant inhibition were observed at 1000 mg/kg. However, histamine-induced scratching behavior was not inhibited by propolis granular A.P.C even at 1000 mg/kg. Propolis ethanol extract at 10 microg/ml or more inhibited histamine release from rat mast cells induced by compound 48/80. In addition, it blocked increased vascular permeability induced by compound 48/80. The inhibitory effect of propolis on scratching behavior induced by compound 48/80 was gradually enhanced by repeated administration, and 500 mg/kg propolis granular A.P.C, which caused no effect through single administration, significantly inhibited scratching behavior after repeated administration for 4 weeks. From these findings, it is assumed that the inhibition of scratching behavior induced by propolis occurs through a mast cell-dependent mechanism.

In vitro permeation through porcine buccal mucosa of caffeic acid phenetyl ester (CAPE) from a topical mucoadhesive gel containing propolis.

Recent studies have shown that propolis has on the oral cavity appreciable antibacterial, antifungal and antiviral actions, as well as anti-inflammatory, anaesthetic and cytostatic properties. In light of these studies, an assessment of the diffusion and permeation of caffeic acid phenetyl ester (CAPE) through porcine buccal mucosa was considered useful as a possible application in the stomatological field. To do so, a mucoadhesive topical gel was prepared to apply to the buccal mucosa. The gel was formulated in such a way as to improve the solubility of the propolis, conducting to an increase of the flux. The mucosal permeation of CAPE from the formulation was evaluated using Franz cells, with porcine buccal mucosa as septum between the formulation (donor compartment) and the receptor phase chamber. The diffusion through the membrane was determined by evaluating the amount of CAPE present in the receiving solution, the flux and the permeation coefficient (at the steady state) in the different formulations at set intervals. Qualitative and quantitative determinations were done by HPLC analysis. From the results, CAPE allowed a high permeability coefficient in comparison to the coefficient of other molecules, as expected from its physical-chemical structure. Moreover, the developed gel improved the CAPE flux approximately 35 times more with respect to an ethanol solution formulated at the same gel concentration. The developed gel was also tested in order to evaluate the mucoadhesive behaviour and comfort in vivo on 10 volunteers in a period of 8 h. The in vivo evaluation of mucoadhesive gel revealed adequate comfort and non-irritancy during the period of study and it was well accepted by the volunteers.