Tocotrienols Provide Radioprotection to Multiple Organ Systems through Complementary Mechanisms of Antioxidant and Signaling Effects.

Tocotrienols have powerful radioprotective properties in multiple organ systems and are promising candidates for development as clinically effective radiation countermeasures. To facilitate their development as clinical radiation countermeasures, it is crucial to understand the mechanisms behind their powerful multi-organ radioprotective properties. In this context, their antioxidant effects are recognized for directly preventing oxidative damage to cellular biomolecules from ionizing radiation. However, there is a growing body of evidence indicating that the radioprotective mechanism of action for tocotrienols extends beyond their antioxidant properties. This raises a new pharmacological paradigm that tocotrienols are uniquely efficacious radioprotectors due to a synergistic combination of antioxidant and other signaling effects. In this review, we have covered the wide range of multi-organ radioprotective effects observed for tocotrienols and the mechanisms underlying it. These radioprotective effects for tocotrienols can be characterized as (1) direct cytoprotective effects, characteristic of the classic antioxidant properties, and (2) other effects that modulate a wide array of critical signaling factors involved in radiation injury.

Analysis of Plant-Plant Interactions Reveals the Presence of Potent Antileukemic Compounds.

A method to identify anticancer compounds in plants was proposed based on the hypothesis that these compounds are primarily present in plants to provide them with an ecological advantage over neighboring plants and other competitors. According to this view, identifying plants that contain compounds that inhibit or interfere with the development of other plant species may facilitate the discovery of novel anticancer agents. The method was developed and tested using Magnolia grandiflora, Gynoxys verrucosa, Picradeniopsis oppositifolia, and Hedyosmum racemosum, which are plant species known to possess compounds with cytotoxic activities. Plant extracts were screened for growth inhibitory activity, and then a thin-layer chromatography bioautography assay was conducted. This located the major antileukemic compounds 1, 2, 4, and 5 in the extracts. Once the active compounds were located, they were extracted and purified, and their structures were determined. The growth inhibitory activity of the purified compounds showed a significant correlation with their antileukemic activity. The proposed approach is rapid, inexpensive, and can easily be implemented in areas of the world with high biodiversity but with less access to advanced facilities and biological assays.

A pharmacokinetic study of morphine-6-O-sulfate in rat plasma and brain.

Morphine-6-O-sulfate (M6S), a polar, zwitterionic sulfate ester of morphine, is a powerful and safe analgesic in several rat models of pain. A sensitive liquid chromatography-tandem mass spectrometry bioanalytical method was developed and validated for the simultaneous determination of M6S and morphine (MOR) in rat plasma and brain after M6S administration. Morphine-d6 was used as internal standard. Multiple reaction monitoring was used for detection and quantitation of M6S, MOR, and morphine-d6 in the turbo ion spray positive mode. The chromatographic separation was carried out on an Alltech Altima C18 column. The analytical method was validated for linearity, precision, accuracy, specificity, and stability over a concentration range of 3-8000 ng/ml in rat plasma and 10-10,000 ng/ml in brain samples for both M6S and MOR. The validated method was applied to determine the PK profile of M6S in plasma after i.v., i.p., and oral dosing in male Sprague-Dawley rats. Rats were administered M6S by i.p. administration (5.6 and 10.0 mg/kg) or orally (10 and 30 mg/kg) and bioavailability compared to an i.v. injection (1 mg/kg) of M6S. The in vivo results indicate that M6S is not a prodrug of morphine, since M6S is not biotransformed into MOR in plasma after either i.p. or oral administration, and MOR was not detected in brain. The bioavailability of M6S was >93% and about 5% after i.p. and oral dosing, respectively. The low oral bioavailability of M6S may be due to poor permeation of the intestinal epithelial membrane. After i.p.-administration, M6S appears to reach brain tissues in low, but significant, concentrations.

Antioxidant Tocols as Radiation Countermeasures (Challenges to be Addressed to Use Tocols as Radiation Countermeasures in Humans).

Radiation countermeasures fall under three categories, radiation protectors, radiation mitigators, and radiation therapeutics. Radiation protectors are agents that are administered before radiation exposure to protect from radiation-induced injuries by numerous mechanisms, including scavenging free radicals that are generated by initial radiochemical events. Radiation mitigators are agents that are administered after the exposure of radiation but before the onset of symptoms by accelerating the recovery and repair from radiation-induced injuries. Whereas radiation therapeutic agents administered after the onset of symptoms act by regenerating the tissues that are injured by radiation. Vitamin E is an antioxidant that neutralizes free radicals generated by radiation exposure by donating H atoms. The vitamin E family consists of eight different vitamers, including four tocopherols and four tocotrienols. Though alpha-tocopherol was extensively studied in the past, tocotrienols have recently gained attention as radiation countermeasures. Despite several studies performed on tocotrienols, there is no clear evidence on the factors that are responsible for their superior radiation protection properties over tocopherols. Their absorption and bioavailability are also not well understood. In this review, we discuss tocopherol's and tocotrienol's efficacy as radiation countermeasures and identify the challenges to be addressed to develop them into radiation countermeasures for human use in the event of radiological emergencies.

Identification of a melampomagnolide B analog as a potential lead molecule for treatment of acute myelogenous leukemia.

A series of carbamate derivatives of the antileukemic sesquiterpene melampomagnolide B (MMB) has been synthesized utilizing a 1,2,4-triazole carbamate conjugate of MMB as an intermediate synthon. Five imidazole- and benzimidazole-carbamate analogs of MMB (8a-8e) were prepared and evaluated for anti-leukemic activity against cultured M9 ENL1 AML cells. All the analogs exhibited improved anti-leukemic activity (EC50=0.90-3.93µM) when compared to parthenolide and the parent sesquiterpene, MMB (EC50=7.0µM and 15.5µM, respectively). The imidazole carbamate analog, 8a (EC50=0.9µM), was 16 times more potent than MMB. The comparative bioavailabilities of 8a and MMB were determined in BALB/c mice following oral dosing of these compounds. It has been demonstrated that the absolute plasma bioavailabilities of MMB and 8a were 6.7±0.8%, and 45.5±2%, respectively. These results indicate that, compared to MMB, the PK parameters for 8a display significantly improved bioavailability and exposure after oral administration. Analog 8a is considered to be a potential clinical candidate for treatment of acute myelogenous leukemia.

Novel Bone-Targeting Agent for Enhanced Delivery of Vancomycin to Bone.

We examined the pharmacokinetic properties of vancomycin conjugated to a bone-targeting agent (BT) with high affinity for hydroxyapatite after systemic intravenous administration. The results confirm enhanced persistence of BT-vancomycin in plasma and enhanced accumulation in bone relative to vancomycin. This suggests that BT-vancomycin may be a potential carrier for the systemic targeted delivery of vancomycin in the treatment of bone infections, potentially reducing the reliance on surgical debridement to achieve the desired therapeutic outcome.

Tocotrienol-Rich Fraction from Rice Bran Demonstrates Potent Radiation Protection Activity.

The vitamin E analogs δ-tocotrienol (DT3) and γ-tocotrienol (GT3) have significant protective and mitigative capacity against the detrimental effects of ionizing radiation (IR). However, the expense of purification limits their potential use. This study examined the tocotrienol-rich fraction of rice bran (TRFRB) isolated from rice bran deodorizer distillate, a rice oil refinement waste product, to determine its protective effects against IR induced oxidative damage and H2O2. Several cell lines were treated with tocotrienols or TRFRB prior to or following exposure to H2O2 or IR. To determine the radioprotective capacity cells were analyzed for morphology, mitochondrial bioenergetics, clonogenic survival, glutathione oxidation, cell cycle, and migration rate. TRFRB displayed similar antioxidant activity compared to pure tocotrienols. Cells pretreated with TRFRB or DT3 exhibited preserved cell morphology and mitochondrial respiration when exposed to H2O2. Oxidized glutathione was decreased in TRFRB treated cells exposed to IR. TRFRB reversed mitochondrial uncoupling and protected cells migration rates following IR exposure. The protective antioxidant capacity of TRFRB treated cells against oxidative injury was similar to that of purified DT3. TRFRB effectively protects normal cells against IR induced injury suggesting that rice bran distillate may be an inexpensive and abundant alternate source.

Molecular dynamics guided design of tocoflexol: a new radioprotectant tocotrienol with enhanced bioavailability.

There is a pressing need to develop safe and effective radioprotector/radiomitigator agents for use in accidental or terrorist-initiated radiological emergencies. Naturally occurring vitamin E family constituents, termed tocols, that include the tocotrienols, are known to have radiation-protection properties. These agents, which work through multiple mechanisms, are promising radioprotectant agents having minimal toxicity. Although α-tocopherol (AT) is the most commonly studied form of vitamin E, the tocotrienols are more potent than AT in providing radioprotection and radiomitigation. Unfortunately, despite their very significant radioprotectant activity, tocotrienols have very short plasma half-lives and require dosing at very high levels to achieve necessary therapeutic benefits. Thus, it would be highly desirable to develop new vitamin E analogues with improved pharmacokinetic properties, specifically increased elimination half-life and increased area under the plasma level versus time curve. The short elimination half-life of the tocotrienols is related to their low affinity for the α-tocopherol transfer protein (ATTP), the protein responsible for maintaining the plasma level of the tocols. Tocotrienols have less affinity for ATTP than does AT, and thus have a longer residence time in the liver, putting them at higher risk for metabolism and biliary excretion. We hypothesized that the low-binding affinity of tocotrienols to ATTP is due to the relatively more rigid tail structure of the tocotrienols in comparison with that of the tocopherols. Therefore, compounds with a more flexible tail would have better binding to ATTP and consequently would have longer elimination half-life and, consequently, an increased exposure to drug, as measured by area under the plasma drug level versus time curve (AUC). This represents an enhanced residence of drug in the systemic circulation. Based on this hypothesis, we developed a new class of vitamin E analogues, the tocoflexols, which maintain the superior bioactivity of the tocotrienols with the potential to achieve the longer half-life and larger AUC of the tocopherols.

Drug dissolution: significance of physicochemical properties and physiological conditions.

Oral bioavailability of a drug is determined by a number of properties, including drug dissolution rate, solubility, intestinal permeability and pre-systemic metabolism. Frequently, the rate limiting step in drug absorption from the gastrointestinal tract is drug release and drug dissolution from the dosage form. Therapeutic agents with aqueous solubilities less than 100µg/ml often present dissolution limitations to absorption. Physicochemical, formulation-related and physiological factors can all influence drug dissolution. In this review, the authors will discuss the important physicochemical properties of a drug and physiological conditions in the gastrointestinal tract that play an important part in drug dissolution and absorption processes and, consequently, the bioavailability of a drug.

Sesquiterpene lactones from Gynoxys verrucosa and their anti-MRSA activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Because of its virulence and antibiotic resistance, Staphylococcus aureus is a more formidable pathogen now than at any time since the pre-antibiotic era. In an effort to identify and develop novel antimicrobial agents with activity against this pathogen, we have examined Gynoxys verrucosa Wedd (Asteraceae), an herb used in traditional medicine in southern Ecuador for the treatment and healing of wounds. MATERIALS AND METHODS: The sesquiterpene lactones leucodine (1) and dehydroleucodine (2) were extracted and purified from the aerial parts of Gynoxys verrucosa, and their structure was elucidated by spectroscopic methods and single-crystal X-ray analysis. The in vitro anti-microbial activity of Gynoxys verrucosa extracts and its purified constituents was determined against six clinical isolates including Staphylococcus aureus and Staphylococcus epidermidis strains with different drug-resistance profiles, using the microtiter broth method. RESULTS: Compound 1 has very low activity, while compound 2 has moderate activity with MIC(50)s between 49 and 195 µg/mL. The extract of Gynoxys verrucosa has weak activity with MIC(50)s between 908 and 3290 µg/mL. CONCLUSIONS: We are reporting the full assignment of the (1)H NMR and (13)C NMR of both compounds, and the crystal structure of compound 2, for the first time. Moreover, the fact that compound 2 has antimicrobial activity and compound 1 does not, demonstrates that the exocyclic conjugated methylene in the lactone ring is essential for the antimicrobial activity of these sesquiterpene lactones. However, the weak activity observed for the plant extracts, does not explain the use of Gynoxys verrucosa in traditional medicine for the treatment of wounds and skin infections.

3D-QSAR, synthesis, and antimicrobial activity of 1-alkylpyridinium compounds as potential agents to improve food safety.

Cetylpyridinium chloride (CPC), an alkylpyridinium compound has been recently approved by the US Food and Drug Administration to reduce bacterial contamination in poultry. Although CPC is very effective and has a very good safety record, its relatively high lipophilicity may limit its use in high fat containing foods such as beef. In this study we present the CoMFA analysis (3D-QSAR) of the antimicrobial activity of 60 N-alkylpyridinium compounds against different bacteria. CoMFA contours showed that the activity is highly influenced by the steric factor. Based in these contours we designed new candidates, which were synthesized and characterized by spectroscopic data. MIC activity over Gram positive and Gram negative microorganisms validated the 3D-QSAR study.

Liquid chromatography determination of residue levels on apples treated with cetylpyridinium chloride.

Cetylpyridinium chloride (CPC) has been found to be effective in reducing microbial contamination in apples. A sensitive and specific HPLC method was developed to determine CPC residues in apples treated with CPC. This method involves ion exchange solid-phase extraction, and the use of stearylpyridinium chloride (SPC) as internal standard. Limit of quantitation, was 0.5 microg/ml of CPC for the apple ethanolic extracts. The observed residues in apple (2.35-4.35 microg/g of apple) were lower than those previously reported for chicken and beef. The method is specific, sensitive, reproducible and accurate.

Elimination of Salmonella Contamination from Poultry Tissues by Cetylpyridinium Chloride Solutions.

The effects of cetylpyridinium chloride (CPC) on the inhibition and reduction of viable Salmonella typhimurium cells were studied. In these experiments skin excised from chicken drumsticks was treated with solutions of CPC. At the CPC concentrations used, the ability of this compound to reduce bacterial contamination was clearly demonstrated. This effect was both CPC concentration- and exposure-time-dependent. A 4.87-log reduction of viable S. typhimurium cells was achieved at a CPC concentration of 4 mg/ml at the treatment time of 3 min. Moreover, CPC was effective in preventing bacterial contamination, as shown by a 4.9-log inhibition of S. typhimurium cell attachment at a CPC concentration of 8 mg/ml and a treatment time of 10 min.