Development of nanoparticles coated with cassava bagasse pectin (Manihot esculenta Crantz) containing ß-carotene for mucoadhesive applications.

Pectin (PC) extracted from a solid residue from cassava roots (Manihot esculenta Crantz) was used to coat nanoparticles (NP) containing ß-carotene (BC) aiming at the gastrointestinal administration of this lipophilic nutraceutical. The NP were prepared by spontaneous emulsification method using food grade components. Pectin-coated NP have been successfully prepared as confirmed by the increased particle size and negative surface charges due to the pectin's anionic nature. NP showed spherical shape and monodisperse distribution, with a mean size of 21.3 nm (polydispersity index (PDI) 0.29) for BC PC T80-NP (nanoparticle with ß-carotene, pectin and Tween 80) and 261.4 nm (PDI 0.1) for BC PC T20-NP (nanoparticle with ß-carotene, pectin and Tween 20). BC was encapsulated at amounts of 530 and 324 µg/ml for BC PC T80-NP and BC PC T20-NP, respectively, with high encapsulation efficiency (> 95%), increasing its antioxidant capacity in vitro, besides no cytotoxic effect. However, only BC PC T20-NP was stable over a 90 days storage period (4°C) and revealed a strong interaction between pectin and mucin. These results suggest that pectin-coated BC PC T20-NP is a promising strategy to improve the bioavailability and permeation of BC for administration through mucosal surfaces.

Determination of l-(+)-bornesitol, the hypotensive constituent of Hancornia speciosa, in rat plasma by LC-MS/MS and its application on a pharmacokinetic study.

Hancornia speciosa is a medicinal plant with proven antihypertensive activity. The cyclitol l-(+)-bornesitol is the main constituent of its leaves and is a potent inhibitor of the angiotensin-converting enzyme. We herein investigated the pharmacokinetic properties of bornesitol administered orally to Wistar rats, as well as bornesitol permeation in Caco-2 cells. Bornesitol was isolated and purified from an ethanol extract of H. speciosa leaves. An ultra-high performance liquid chromatography coupled with electrospray ionization mass spectrometry (UPLC-ESI-MS/MS) method was developed and validated to quantify bornesitol in rat plasma based on Multiple Reaction Monitoring, using pentaerythritol as an internal standard. Pharmacokinetics was evaluated by the administration of single doses via intravenous in bolus (3 mg/kg) and gavage (3, 15 and 25 mg/kg). Bornesitol permeation was assayed in a transwell Caco-2 cells model, tested alone, or combined with rutin, or as a constituent of H. speciosa extract, using a developed and validated UPLC-ESI-MS/MS method. All assayed validation parameters (selectivity, residual effect, matrix effect, linearity, precision, accuracy and stability of analyte in plasma and solution) for the bioanalytical method met the acceptance criteria established by regulatory guidelines. Bornestiol reached peak plasma concentration within approximately 60 min after oral administration with a half-life ranging from 72.15 min to 123.69 min. The peak concentration and area under the concentration-time curve of bornesitol did not rise proportionally with the increasing doses, suggesting a non-linear pharmacokinetics in rats and the oral bioavailability ranged from 28.5%-59.3%. Bornesitol showed low permeability in Caco-2 cells, but the permeability apparently increased when it was administered either combined with rutin or as a constituent of H. speciosa extract. In conclusion, bornesitol was rapidly absorbed after a single oral administration to rats and followed a non-linear pharmacokinetics. The obtained data will be useful to guide further pre-clinical development of bornesitol-containing herbal preparations of H. speciosa as an antihypertensive agent.

Antiviral Potential of Spondias mombin L. Leaves Extract Against Herpes Simplex Virus Type-1 Replication Using In Vitro and In Silico Approaches.

Spondias mobin leaves have been traditionally used for treating cold sores. The study investigated the mechanism of antiherpes action of S. mombin extract, fractions, and geraniin. Different concentrations of samples were used to evaluate the in vitro antiherpes activity (anti-HSV-1) in virucidal, post-infection, attachment, and penetration assays. The mechanism of action of geraniin was investigated considering the glycoproteins gB and gD of HSV-1 surface as potential molecular targets. Molecular docking simulations were carried out for both in order to determine the possible binding mode position of geraniin at the activity sites. The binding mode position was posteriorly optimized considering the flexibility of the glycoproteins. The chemical analysis of samples was performed by LC-MS and revealed the presence of 22 substances, which are hydrolysable tannins, O-glycosylated flavonoids, phenolic acids, and a carbohydrate. The extract, tannin-rich fraction and geraniin showed important in vitro virucidal activity through blocking viral attachment but showed no relevant inhibition of viral penetration. The in silico approaches demonstrated a high number of potential strong intermolecular interactions as hydrogen bonds between geraniin and the activity site of the glycoproteins, particularly the glycoprotein gB. In silico experiments indicated that geraniin is at least partially responsible for the anti-herpes activity through interaction with the viral surface glycoprotein gB, which is responsible for viral adsorption. These results highlight the therapeutic potential of S. mombin anti-herpes treatment and provides support for its traditional purposes. However, further studies are required to validate the antiviral activities in vivo, as well as efficacy in humans.

Blackberry extract improves behavioral and neurochemical dysfunctions in a ketamine-induced rat model of mania.

Bipolar disorder is a chronic mood disorder characterized by episodes of mania and depression. The aim of this study was to investigate the effects of blackberry extract on behavioral parameters, oxidative stress and inflammatory markers in a ketamine-induced model of mania. Animals were pretreated with extract (200 mg/kg, once a day for 14 days), lithium chloride (45 mg/kg, twice a day for 14 days), or vehicle. Between the 8th and 14th days, the animals received an injection of ketamine (25 mg/kg) or vehicle. On the 15th day, thirty minutes after ketamine administration, the animals' locomotion was assessed using open-field apparatus. After the experiments, the animals were euthanized and cerebral structures were removed for neurochemical analyses. The results showed that ketamine treatment induced hyperlocomotion and oxidative damage in the cerebral cortex, hippocampus and striatum. In contrast, pretreatment with the extract or lithium was able to prevent hyperlocomotion and oxidative damage in the cerebral cortex, hippocampus, and striatum. In addition, IL-6 and IL-10 levels were increased by ketamine, while the extract prevented these effects in the cerebral cortex. Pretreatment with the extract was also effective in decreasing IL-6 and increasing the level of IL-10 in the striatum. In summary, our findings suggest that blackberry consumption could help prevent or reduce manic episodes, since this extract have demonstrated neuroprotective properties as well as antioxidant and anti-inflammatory effects in the ketamine-induced mania model.

Chemical Constituents and Pharmacology properties of Aristolochia triangularis: a south brazilian highly-consumed botanical with multiple bioactivities.

Aristolochia triangularis Cham., is one of the most frequently used medicinal plant in Southern Brazil. Preparations containing the leaves and/or stems are traditionally used as anti-inflammatory, diuretic, as well as antidote against snakebites. This study screened A. triangularis extracts, fractions and isolated compounds for different bioactivities. A weak antiproliferative activity against human lung cancer cell line (A549) was observed only for chloroform fraction obtained from stems (CFstems - CC50: 2.93 µg/mL). Also, a moderate antimicrobial activity against Staphylococcus aureus was detected just for chloroform fraction obtained from leaves (CFleaves -13-16 mm inhibition zone). Additionally, two semi-purified fractions (CFstems-4 and CFleaves-4) selectively inhibited HSV-1 replication (IC50 values of 0.40 and 2.61 µg/mL, respectively), while only CFleaves showed promising results against Leishmania amazonensis. Fractionation of extracts resulted in the isolation of one neolignan (-) cubebin and one lignan (+) galbacin. However, these compounds are not responsible for the in vitro bioactivities herein detected. The presence of aristolochic acid I and aristolochic acid II in the crude ethanol extract of stems (CEEstems) and leaves (CEEleaves) was also investigated. The HPLC analysis of these extracts did not display any peak with retention time or UV spectra comparable to aristolochic acids I and II.

New 99mTc-Labeled Digitoxigenin Derivative for Cancer Cell Identification.

In recent years, cardiac glycosides (CGs) have been investigated as potential antiviral and anticancer drugs. Digitoxigenin (DIG) and other CGs have been shown to bind and inhibit Na+/K+-adenosinetriphosphatase (ATPase). Tumor cells show a higher expression rate of the Na+/K+-ATPase protein or a stronger affinity towards the binding of CGs and are therefore more prone to CGs than non-tumor cells. Cancer imaging techniques using radiotracers targeted at specific receptors have yielded successful results. Technetium-99m (99mTc) is one of the radionuclides of choice to radiolabel pharmaceuticals because of its favorable physical and chemical properties along with reasonable costs. Herein, we describe a new Na+/K+-ATPase targeting radiotracer consisting of digitoxigenin and diethylenetriaminepentaacetic acid (DTPA), a bifunctional chelating ligand used to prepare 99mTc-labeled complexes, and its evaluation as an imaging probe. We report the synthesis and characterization of the radiolabeled compound including stability tests, blood clearance, and biodistribution in healthy mice. Additionally, we investigated the binding of the compound to A549 human non-small-cell lung cancer cells and the inhibition of the Na+/K+-ATPase by the labeled compound in vitro. The 99mTc-labeled DTPA-digitoxigenin (99mTc-DTPA-DIG) compound displayed high stability in vitro and in vivo, a fast renal excretion, and a specific binding towards A549 cancer cells in comparison to non-tumor cells. Therefore, 99mTc-DTPA-DIG could potentially be used for non-invasive visualization of tumor lesions by means of scintigraphic imaging.

Nanoemulsion-Loaded Hydrogels for Topical Administration of Pentyl Gallate.

The aim of this study is to describe the development of nanoemulsion-loaded hydrogels to deliver pentyl gallate (PG), a gallic acid n-alkyl ester, through the skin. PG is an antioxidant agent; however, it seems to be a promising agent for herpis labialis treatment. Aristoflex AVC® and chitosan were used as gelling agents for nanoemulsion thickening. The developed formulations presented suitable PG content (94.4-100.3% w/w), nanometric droplet sizes (162-297 nm), high zeta potentials, and a non-Newtonian pseudoplastic behavior. Both vehicles neither enhanced PG penetration nor delayed its release from the nanoemulsion. Formulations remained physically stable at 8°C during 3 months of storage.

Coumestrol/hydroxypropyl-ß-cyclodextrin association incorporated in hydroxypropyl methylcellulose hydrogel exhibits wound healing effect: in vitro and in vivo study.

Several beneficial effects on the skin have been reported for coumestrol (COU), such as protection against photoaging and improvement of skin elasticity and thickness in postmenopausal women. However no reports on the effect of COU on wound healing were found. Nevertheless, COU has low aqueous solubility, which is a crucial limitation for biological tests. The present study was designed as a two-step experiment to evaluate the wound healing effect of COU. First, we used fibroblasts and the experimental in vitro artificial wound model, scratch assay, to compare the effects of COU free, dissolved in dimethyl sulfoxide (DMSO) or Dulbecco's modified Eagle's medium (DMEM), or associated with hydroxypropyl-ß-cyclodextrin (HPßCD). The 50 µM (66.1%) and 10 µM (56.3%) COU/HPßCD association induced cell proliferation and migration in inflicted wounds. Subsequently, the in vivo wound healing experimental model (Wistar rats) revealed that COU/HPßCD incorporated into hypromellose (HPMC) hydrogel had similar efficacy in wound healing in comparison to the positive control (Dersani®), with the advantage that 50% wound healing was achieved within a shorter period. In summary, the results successfully demonstrated, for the first time, the wound healing effect of COU/HPßCD incorporated into HPMC hydrogel and describe the feasibility of the biological tests with the use of HPßCD instead DMSO.

Cardiac Glycoside Glucoevatromonoside Induces Cancer Type-Specific Cell Death.

Cardiac glycosides (CGs) are natural compounds used traditionally to treat congestive heart diseases. Recent investigations repositioned CGs as potential anticancer agents. To discover novel cytotoxic CG scaffolds, we selected the cardenolide glucoevatromonoside (GEV) out of 46 CGs for its low nanomolar anti-lung cancer activity. GEV presented reduced toxicity toward non-cancerous cell types (lung MRC-5 and PBMC) and high-affinity binding to the Na+/K+-ATPase α subunit, assessed by computational docking. GEV-induced cell death was caspase-independent, as investigated by a multiparametric approach, and culminates in severe morphological alterations in A549 cells, monitored by transmission electron microscopy, live cell imaging and flow cytometry. This non-canonical cell death was not preceded or accompanied by exacerbation of autophagy. In the presence of GEV, markers of autophagic flux (e.g. LC3I-II conversion) were impacted, even in presence of bafilomycin A1. Cell death induction remained unaffected by calpain, cathepsin, parthanatos, or necroptosis inhibitors. Interestingly, GEV triggered caspase-dependent apoptosis in U937 acute myeloid leukemia cells, witnessing cancer-type specific cell death induction. Differential cell cycle modulation by this CG led to a G2/M arrest, cyclin B1 and p53 downregulation in A549, but not in U937 cells. We further extended the anti-cancer potential of GEV to 3D cell culture using clonogenic and spheroid formation assays and validated our findings in vivo by zebrafish xenografts. Altogether, GEV shows an interesting anticancer profile with the ability to exert cytotoxic effects via induction of different cell death modalities.

Cytotoxic and cytostatic effects of digitoxigenin monodigitoxoside (DGX) in human lung cancer cells and its link to Na,K-ATPase.

Cardiac glycosides (CGs) are natural compounds widely used to treat several cardiac conditions and more recently have been recognized as potential antitumor agents. They are known as Na,K-ATPases ligands, which is a promising drug target in cancer. In this study, the short and long-lasting cytotoxic effects of the natural cardenolide digitoxigenin monodigitoxoside (DGX) were evaluated against two non-small cell lung cancer lines (A549 and H460 cells). It was found that DGX induced cytotoxic effects in both cells and the apoptotic effects were more pronounced on H460 cells. In long-term analysis, using the clonogenic and the cumulative population doubling (CPD) assays, DGX showed a reduction of cell survival, after 15days without re-treatment. To better understand DGX effects in A549 cells, several assays were conducted. In cell cycle analysis, DGX caused an arrest in S and G2/M phases. This compound also increased the number of cells in subG1 phase in a concentration- and time-dependent manner. The presence of ß-galactosidase positive cells, large nucleus and flattened cells indicated senescence. Additionally, DGX inhibited Na,K-ATPase activity in A549 cells, as well as in purified pig kidney and in human red blood cell membrane preparations, at nanomolar range. Moreover, results of molecular docking showed that DGX binds with high efficiency (-11.4Kcal/mol) to the Na,K-ATPase (PDB:4HYT). Taken together, our results highlight the potent effects of DGX both in A549 and H460 cells, and disclose its link with Na,K-ATPase inhibition.

Pharmacokinetics of Botanical Drugs and Plant Extracts.

BACKGROUND: Botanical drugs contain plant extracts, which are complex mixtures of compounds. As with conventional drugs, it is necessary to validate their efficacy and safety through preclinical and clinical studies. However, pharmacokinetic studies for active constituents or characteristic markers in botanical drugs are rare. OBJECTIVE: The objective of this review was to investigate the global state of the art in pharmacokinetic studies of active ingredients present in plant extracts and botanical drugs. A review of pharmacokinetics studies of chemical constituents of plant extracts and botanical drugs was performed, with a total of 135 studies published between January 2004 and February 2015 available in recognized scientific databases. Botanical preparations were mainly found in the form of aqueous extracts of roots and rhizomes. The most widely studied species was Salvia miltiorrhiza Bunge, and the compound most frequently used as a pharmacokinetic marker was berberine. CONCLUSION: Most studies were performed using the Sprague Dawley rat model, and the preparations were mainly administered orally in a single dose. Quantification of plasma concentrations of pharmacokinetic markers was performed mainly by liquid-liquid extraction, followed by high performance liquid chromatography coupled to mass spectrometry detector. In conclusion, in recent years there has been an increasing interest among researchers worldwide in the study of pharmacokinetics of bioactive compounds in botanical drugs and plant extracts, especially those from the Traditional Chinese Medicine.

Production of the Cytotoxic Cardenolide Glucoevatromonoside by Semisynthesis and Biotransformation of Evatromonoside by a Digitalis lanata Cell Culture.

Recent studies demonstrate that cardiac glycosides, known to inhibit Na+/K+-ATPase in humans, have increased susceptibility to cancer cells that can be used in tumor therapy. One of the most promising candidates identified so far is glucoevatromonoside, which can be isolated from the endangered species Digitalis mariana ssp. heywoodii. Due to its complex structure, glucoevatromonoside cannot be obtained economically by total chemical synthesis. Here we describe two methods for glucoevatromonoside production, both using evatromonoside obtained by chemical degradation of digitoxin as the precursor. 1) Catalyst-controlled, regioselective glycosylation of evatromonoside to glucoevatromonoside using 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl bromide as the sugar donor and 2-aminoethyldiphenylborinate as the catalyst resulted in an overall 30 % yield. 2) Biotransformation of evatromonoside using Digitalis lanata plant cell suspension cultures was less efficient and resulted only in overall 18 % pure product. Structural proof of products has been provided by extensive NMR data. Glucoevatromonoside and its non-natural 1-3 linked isomer neo-glucoevatromonoside obtained by semisynthesis were evaluated against renal cell carcinoma and prostate cancer cell lines.

Cytotoxic effects of the cardenolide convallatoxin and its Na,K-ATPase regulation.

Cardenolides are cardiac glycosides, mostly obtained from natural sources. They are well known for their inhibitory action on the Na,K-ATPase, an effect that regulates cardiovascular alterations such as congestive heart failure and atrial arrhythmias. In recent years, they have also sparked new interest in their anticancer potential. In the present study, the cytotoxic effects of the natural cardenolide convallatoxin (CON) were evaluated on non-small cell lung cancer (A549 cells). It was found that CON induced cytostatic and cytotoxic effects in A549 cells, showing essentially apoptotic cell death, as detected by annexin V-propidium iodide double-staining, as well as changes in cell form. In addition, it prompted cell cycle arrest in G2/M and reduced cyclin B1 expression. This compound also increased the number of cells in subG1 in a concentration- and time-dependent manner. At a long term, the reduction of cumulative population doubling was shown along with an increase of ß-galactosidase positive cells and larger nucleus, indicative of senescence. Subsequently, CON inhibited the Na,K-ATPase in A549 cells at nM concentrations. Interestingly, at the same concentrations, CON was unable to directly inhibit the Na,K-ATPase, either in pig kidney or in red blood cells. Additionally, results of docking calculations showed that CON binds with high efficiency to the Na,K-ATPase. Taken together, our data highlight the potent anticancer effects of CON in A549 cells, and their possible link with non-classical inhibition of Na,K-ATPase.

Solid dispersions enhance solubility, dissolution, and permeability of thalidomide.

Thalidomide (THD) is a BCS class II drug with renewed and growing therapeutic applicability. Along with the low aqueous solubility, additional poor biopharmaceutical properties of the drug, i.e. chemical instability, high crystallinity, and polymorphism, lead to a slow and variable oral absorption. In this view, we developed solid dispersions (SDs) containing THD dispersed in different self-emulsifying carriers aiming at an enhanced absorption profile for the drug. THD was dispersed in lauroyl macrogol-32 glycerides (Gelucire® 44/14) and α-tocopherol polyethylene glycol succinate (Kolliphor® TPGS), in the presence or absence of the precipitation inhibitor polyvinylpyrrolidone K30 (PVP K30), by means of the solvent method. Physicochemical analysis revealed the formation of semicrystalline SDs. X-ray diffraction and infrared spectroscopy analyses suggest that the remaining crystalline fraction of the drug in the SDs did not undergo polymorphic transition. The impact of the solubility-enhancing formulations on the THD biopharmaceutical properties was evaluated by several in vitro techniques. The developed SDs were able to increase the apparent solubility of the drug (up to 2-3x the equilibrium solubility) for a least 4 h. Dissolution experiments (paddle method, 75 rpm) in different pHs showed that around 80% of drug dissolved after 120 min (versus 40% of pure crystalline drug). Additionally, we demonstrated the enhanced solubility obtained via SDs could be translated into increased flux in a parallel artificial membrane permeability assay (PAMPA). In summary, the results demonstrate that SDs could be considered an interesting and unexplored strategy to improve the biopharmaceutical properties of THD, since SDs of this important drug have yet to be reported.

Enzymatic synthesis of ascorbyl ester derived from linoleic acid.

Antioxidants are substances that defend cells against damage, kidnapping and destroying free radicals. They have been largely used in the food industry due the possibility to control the oxidation process, aimed to increase shelf life. Thus, esterification reaction to obtain ascorbyl linoleate catalyzed by Novozym 435 lipase assisted by ultrasound bath was investigated. In this work, molecular sieve (4 Å) was added to the reaction medium to remove the water formed during the esterification reaction to improve the process performance. According to the results, ascorbyl linoleate production up to 90 % was reached after 1 h of reaction time carried out using ultrasound bath, 1:9 molar ratio of substrates L-ascorbic acid to linoleic acid, 20 mL of tert-butanol as organic solvent, 5 wt% of Novozym 435 lipase, 10 wt% of molecular sieve at 70 °C.

Feasibility study with 99mTc-HYNIC-ßAla-Bombesin(7-14) as an agent to early visualization of lung tumour cells in nude mice.

AIM: More sensitive and accurate imaging approaches for early detection and therapy monitoring of lung tumours are needed to ameliorate prognosis and outcome. Lung tumours are known to overexpress receptors for bombesin-like peptides. However, thus far, no study has demonstrated the potential role of bombesin-like peptides in identifying A549 lung tumour cells in xenograft animal models. Thus, we evaluate the feasibility of Tc-HYNIC-ßAla-Bombesin(7-14) as an imaging probe in lung cancer. METHODS AND RESULTS: Xenograft lung tumours were implanted in nude mice and evaluated by histopathological analysis. Tumours were easily visualized by Tc-HYNIC-ßAla-Bombesin(7-14) within 30 days after inoculation of the A549 cell line into mice. Scintigraphic images showed high tumour-to-background ratio. DISCUSSION: The data obtained in this study indicate that Tc-HYNIC-ßAla-Bombesin(7-14) may be useful as an imaging probe to detect A549 lung cancer cells. To our knowledge, this is the first time that this specific radiocompound has been used to visualize non-small-cell lung cancer A549 in mice. Further translational research in humans is required to establish the potential role of this radiocompound in clinical practice.

Inhibition of cell proliferation, invasion and migration by the cardenolides digitoxigenin monodigitoxoside and convallatoxin in human lung cancer cell line.

Cardiac glycosides consist of a large family of naturally derived compounds that are clinically used to treat congestive heart failure, and also present anticancer properties. In this study, the cytotoxic effects of two cardenolides, digitoxigenin monodigitoxoside (DGX) and convallatoxin (CON) were screened in four human tumour cell lines. Both compounds showed anti-proliferative effects in all tumour cells, at nanomolar concentrations. Since the human lung cancer cell line A549 was the most sensitive, we investigated the anti-proliferative, anti-migratory and anti-invasive effects of these cardenolides. DGX and CON reduced A549 cell migration, being able to reduce more than 90% of cell invasion. Their effects on the expression of key regulators of metastatic mechanism showed decreased levels of MMP-2, MMP-9 and p-FAK. Both compounds also presented low toxicity for healthy cells. Finally, this work provides the first insights into the effects of these cardenolides on key steps of lung cancer metastasis.

In vitro and in vivo antitumor activity of a novel semisynthetic derivative of cucurbitacin B.

Lung cancer is the most deadly type of cancer in humans, with non-small-cell lung cancer (NSCLC) being the most frequent and aggressive type of lung cancer showing high resistance to radiation and chemotherapy. Despite the outstanding progress made in anti-tumor therapy, discovering effective anti-tumor drugs is still a challenging task. Here we describe a new semisynthetic derivative of cucurbitacin B (DACE) as a potent inhibitor of NSCLC cell proliferation. DACE arrested the cell cycle of lung epithelial cells at the G2/M phase and induced cell apoptosis by interfering with EGFR activation and its downstream signaling, including AKT, ERK, and STAT3. Consistent with our in vitro studies, intraperitoneal application of DACE significantly suppressed the growth of mouse NSCLC that arises from type II alveolar pneumocytes due to constitutive expression of a human oncogenic c-RAF kinase (c-RAF-1-BxB) transgene in these cells. Taken together, these findings suggest that DACE is a promising lead compound for the development of an anti-lung-cancer drug.

Enhancing the buccal mucosal delivery of peptide and protein therapeutics.

With continuing advances in biotechnology and genetic engineering, there has been a dramatic increase in the availability of new biomacromolecules, such as peptides and proteins that have the potential to ameliorate the symptoms of many poorly-treated diseases. Although most of these macromolecular therapeutics exhibit high potency, their large molecular mass, susceptibility to enzymatic degradation, immunogenicity and tendency to undergo aggregation, adsorption, and denaturation have limited their ability to be administered via the traditional oral route. As a result, alternative noninvasive routes have been investigated for the systemic delivery of these macromolecules, one of which is the buccal mucosa. The buccal mucosa offers a number of advantages over the oral route, making it attractive for the delivery of peptides and proteins. However, the buccal mucosa still exhibits some permeability-limiting properties, and therefore various methods have been explored to enhance the delivery of macromolecules via this route, including the use of chemical penetration enhancers, physical methods, particulate systems and mucoadhesive formulations. The incorporation of anti-aggregating agents in buccal formulations also appears to show promise in other mucosal delivery systems, but has not yet been considered for buccal mucosal drug delivery. This review provides an update on recent approaches that have shown promise in enhancing the buccal mucosal transport of macromolecules, with a major focus on proteins and peptides.

Polyoxygenated steroids from the octocoral Leptogorgia punicea and in vitro evaluation of their cytotoxic activity.

Five new polyoxygenated marine steroids-punicinols A-E (1-5)-were isolated from the gorgonian Leptogorgia punicea and characterized by spectroscopic methods (IR, MS, 1H, 13C and 2-D NMR). The five compounds induced in vitro cytotoxic effects against lung cancer A549 cells, while punicinols A and B were the most active, with IC50 values of 9.7 µM and 9.6 µM, respectively. The synergistic effects of these compounds with paclitaxel, as well as their effects on cell cycle distribution and their performance in the clonogenic assay, were also evaluated. Both compounds demonstrated significant synergistic effects with paclitaxel.