Safety Assessment of Lecithin and Other Phosphoglycerides as Used in Cosmetics.

The phosphoglycerides considered in this safety assessment are reported to function primarily as skin and hair conditioning agents, emulsifying agents, and surfactants in cosmetic products and are used up to a maximum reported concentration of 50%. Although phospholipids exert physiologic effects, these are not reproduced by application of phospholipid ingredients to the skin. Given the possibility that Lecithin may be derived from animal sources, it should be noted that the Food and Drug Administration does not permit the use of ingredients made from bovine specified risk materials in cosmetic products. The Expert Panel for Cosmetic Ingredient Safety concluded that the 17 phosphoglycerides are safe in the present practices of use and concentration in cosmetics, as described in this safety assessment.

Pharmacokinetics of Phosphatidylethanol 16:0/20:4 in Human Blood After Alcohol Intake.

BACKGROUND: The purpose of this study was to characterize the pharmacokinetics of the phosphatidylethanol (PEth) 16:0/20:4 homolog in uncoagulated human blood samples taken from 18 participants in a clinical laboratory setting after consumption of 2 standard doses of ethanol (EtOH). METHODS: Male and female participants received either 0.4 or 0.8 g/kg oral doses of EtOH during a 15-minute period. Blood samples were collected before and throughout 6 hours immediately after alcohol administration and then again at days 2, 4, 7, 11, and 14 of the follow-up period. PEth 16:0/20:4 levels were quantified by high-performance liquid chromatography with tandem mass spectrometry detection. RESULTS: (i) The increase in PEth 16:0/20:4 from baseline to maximum concentration was less than that of PEth 16:0/18:1 or PEth 16:0/18:2 homologs during the 6-hour period after EtOH administration; (ii) the mean half-life of PEth 16:0/20:4 was 2.1 ± 3 (SD) days, which was shorter than the mean half-life of either PEth 16:0/18:1 or PEth 16:0/18:2, 7.6 ± 3 (SD) or 6.8 ± 4 (SD) days, respectively. CONCLUSIONS: The pharmacokinetics of PEth 16:0/20:4 in whole blood samples is detectable after alcohol consumption and differs in amount synthesized and rate of elimination versus PEth 16:0/18:1 and 16:0/18:2. Measuring the concentrations of these 3 homologs has the potential to provide more information about the amount and time frame of alcohol consumption than any one alone.

Headgroup-Inversed Liposomes: Biointerfaces, Supported Bilayers and Applications.

Phospholipids are a major component of the cell membrane. In most natural phospholipids, the phosphate acts as a bridge, connecting the other portion of the polar headgroup with the hydrophobic tails. Such bridging phosphate is chemically quite inert. Synthetic lipids inversing the headgroup polarity of phosphocholine (PC) have been recently reported, and these are named CP lipids with a terminal phosphate, or CPe with the terminal phosphate capped by an ethyl group. This Feature Article summarizes the properties and applications of such inversed lipids. First, CPe liposomes were found to be highly resistant to protein adsorption with an even longer blood circulation time than PC liposomes, allowing for enhanced accumulation in tumor sites. CPe liposomes do not interact with PC liposomes either, and this observation was different from that reported using CP polymers, which adhere strongly to cells. Second, CP liposomes interact strongly with many metal oxide nanoparticles (but not silica) forming supported lipid bilayers, while PC liposomes only form supported bilayers on silica. Finally, CP liposomes are good metal ligands based on their exposed terminal phosphate. Zn2+ binds to CP liposomes so strongly that Zn2+ sandwiched multilayered lipid structures were observed. Aside from these fundamental aspects, the potential applications of these headgroup-inversed lipids in drug delivery and biosensor development have also been described, which in turn has promoted fundamental biointerface insights.

Differences in the Synthesis and Elimination of Phosphatidylethanol 16:0/18:1 and 16:0/18:2 After Acute Doses of Alcohol.

BACKGROUND: The purpose of this study was to examine the synthesis and elimination of phosphatidylethanol (PEth) 16:0/18:1 and 16:0/18:2 following the consumption of alcohol among 56 light and heavy drinkers. METHODS: A transdermal alcohol monitor was used to promote alcohol absence 7 days prior, and 14 days after, alcohol consumption in the laboratory. Participants consumed a 0.4 or 0.8 g/kg dose of alcohol in 15 minutes. Blood and breath samples were collected before, at various times up to 360 minutes postconsumption, and 2, 4, 7, 11, and 14 days after alcohol consumption. Initial rates of PEth synthesis, 360 minutes area under the PEth pharmacokinetic curves (AUCs), and elimination half-lives were determined. RESULTS: (i) Nonzero PEth levels were observed before alcohol dosing for most participants, despite 7 days of alcohol use monitoring; (ii) 0.4 and 0.8 g/kg doses of alcohol produced proportional increases in PEth levels in all but 1 participant; (iii) the initial rate of synthesis of both PEth homologues did not differ between the 2 doses, but was greater for PEth 16:0/18:2 than PEth 16:0/18:1 at both doses; (iv) the mean AUC of both PEth homologues was higher at 0.8 g/kg than at 0.4 g/kg; (v) the mean AUC of 16:0/18:2 was greater than that of PEth 16:0/18:1 at both alcohol doses; (vi) the mean half-life of PEth 16:0/18:1 was longer than that of PEth 16:0/18:2 (7.8 ± 3.3 [SD] days and 6.4 ± 5.0 [SD] days, respectively); and (vii) there were no sex differences in PEth 16:0/18:1 or 16:0/18:2 pharmacokinetics. CONCLUSIONS: The results of this study support the use of PEth 16:0/18:1 and 16:0/18:2 as biomarkers for alcohol consumption. Because of consistent pharmacokinetic differences, the levels of these 2 PEth homologues may provide more information regarding the quantity and recentness of alcohol consumption than either alone.

Pharmacokinetics and pharmacodynamics of intravesical and intravenous TMX-101 and TMX-202 in a F344 rat model.

OBJECTIVES: To evaluate and compare the pharmacokinetic and pharmacodynamic properties of 2 investigational Toll-like receptor 7 agonists, TMX-101, and TMX-202 after intravenous and intravesical administration in a rat model. TLR-7 agonists are successfully used as topical treatment for various (pre)malignant skin lesions and are now under investigation as intravesical therapy for non-muscle-invasive bladder cancer. METHODS: Rats received an intravesical instillation with TMX-101, TMX-202, or vehicle. Additionally 2 groups of rats received an intravenous injection with TMX-101 or TMX-202. Blood sampling was performed at different time points, including pre-exposure and postexposure to determine the plasma concentrations of study drugs for pharmacokinetic and pharmacodynamic analyses and to determine the plasma concentrations of cytokines (IL-2, IL-6, and TNF-α). RESULTS: We observed no signs of toxicity after intravesical or intravenous administration. There was a limited dose dependent systemic uptake of TMX-101 and TMX-202 after intravesical administration. The systemic uptake of TMX-202 after intravesical instillation was 25 times lower compared to TMX-101. CONCLUSIONS: This in vivo study confirms the safety of intravesical TMX-101 and TMX-202 administration, with TMX-202 showing lower systemic uptake. TMX-202 has a larger molecule-mass compared to TMX-101, and it may therefore have a favorable safety profile when treating patients with non-muscle-invasive bladder cancer intravesically.

Characterization of the Pharmacokinetics of Phosphatidylethanol 16:0/18:1 and 16:0/18:2 in Human Whole Blood After Alcohol Consumption in a Clinical Laboratory Study.

BACKGROUND: The purpose of this study was to characterize the pharmacokinetics of 2 homologues of phosphatidylethanol (PEth) and their combined total in uncoagulated, whole blood samples taken from participants in a human clinical laboratory study after consumption of low doses of ethanol (EtOH). METHODS: As part of a larger study, 14 male and 13 female participants received either 0.25 or 0.50 g/kg oral doses of EtOH during a 15-minute period. Blood samples were collected before and throughout 6 hours after each EtOH dose on the day of consumption and then every 3 days during the next 14 days. PEth 16:0/18:1 and PEth 16:0/18:2 levels were quantified in blood samples by HPLC/MS/MS and reported separately or as their combined total (combined PEth). Breath alcohol concentrations (BrACs) were measured concurrently with each blood collection. Transdermal alcohol concentrations were measured every 30 minutes during the entire 22-day study to confirm the absence of drinking during a 7-day period before and the 14-day period after EtOH consumption. RESULTS: (i) Single doses of 0.25 and 0.50 g EtOH/kg produced proportional increases in BrAC and combined PEth levels of all participants; (ii) the areas under the curve (AUCs) for each participant's BrAC levels during the 6-hour period after EtOH administration were correlated with AUCs of cPEth (calculated as the AUC of the increase above baseline for combined PEth); (iii) the mean half-life of combined PEth, determined during the 14-day period after EtOH consumption, was 4.6 ± 3.5 (SD) days (range: 1.0 to 13.1 days). CONCLUSIONS: Combined PEth is a sensitive biomarker for the identification of relatively low levels of EtOH consumption. The measurement of these 2 homologues may provide additional sensitivity to identify low levels of drinking.

[Transdermal delivery of insulin]. / Transdermal'nye sistemy vvedeniia insulina.

A possibility is studied of the transdermal delivery of insulin by using a mixture of synthetic analogues of phosphoglycerides (SAP), as a potential activator of hormone diffusion, through the skin. Experimentally in vitro, it was proven that the diffusion of insulin through the skin of two types of transdermal therapeutic form (TTF)--matrix-type and matrix-hydrogel-type--is possible only in presence of activator SAP-M-99. The detected optimal composition of insulin matrix TTF with the area of 40 sq cm enabled a trandermal hormone diffusion speed of 0.26 UNITS/h, which is compatible with the secretion of insulin by the pancreas of an adult (0.25-1.5 UNITS/h). A change-over for the matrix-hydrogel system of insulin delivery based on a 40 sq cm collagenous sponge enabled to increase the insulin diffusion up to 0.54 UNITS/h.

Nonviral vector for efficient gene transfer to human ovarian adenocarcinoma cells.

OBJECTIVE: Various strategies have been attempted to design efficient protocols for ovarian cancer gene therapy but there has been little progress in their clinical application. In this study, we formulated and evaluated a new cationic liposome prepared with dioleoyltrimethylaminopropane (DOTAP), 1,2-dioleoyl-3-phosphophatidylethanolamine (DOPE), and cholesterol (Chol) (DDC) for plasmid DNA transfer into ovarian cancer cells. METHOD: The DDC liposome was prepared by mixing the DOTAP:DOPE:Cholin a 1:0.7:0.3 molar ratio using the extrusion method. Plasmid DNA (pEGFP-C1) and DDC were complexed at various weight ratios to find the optimum condition and the percentage of transfected cells was determined by selecting a green fluorescence protein (GFP) expressing cells in flow cytometry. The transfection efficiency of the DDC liposome was compared with 3[N-(N,N-dimethylaminoethylene) carbamoyl] cholesterol (DC-Chol)/DOPE liposome and commercially available lifopectin. RESULTS: The optimal transfection of plasmid DNA was achieved at a 1:4 (w/w) ratio of DDC to DNA. The DDC/DNA complex exhibited higher transfection efficiency in human ovarian cancer cells (OVCAR-3 and SK-OV-3 cells) compared to that in other types of cell lines (NCI-NIH:522 and HepG2 cells). Flow cytometric analysis revealed that the DDC/DNA complex exhibited an over fourfold increase in GFP expression levels compared with DC-Chol/DOPE or lipofectin in OVCAR-3 cells. This result was further confirmed by confocal microscopy and RT-PCR analysis. CONCLUSION: These results suggest that our newly formulated cationic liposome (DDC) appears to be a promising nonviral vector for treating ovarian adenocarcinoma because of its selective high gene transfer ability in ovarian cancer cells.

Cationic polymer and lipids augment adenovirus-mediated gene transfer to cerebral arteries in vivo.

Adenovirus-mediated gene transfer to blood vessels is relatively inefficient because binding of adenovirus to vessels is limited. The authors have reported that incorporation of cationic polymer and lipids with adenovirus augments gene transfer to blood vessels ex vivo. In this study, the authors determined whether complexes of adenovirus and cations improve efficiency of gene transfer in vivo. Poly-L-lysine, lipofectamine, or lipofectin was complexed with adenovirus encoding beta-galactosidase. Optimum ratios of the cations per adenovirus were determined by gene transfer to fibroblasts. After injection of the adenovirus into the cisterna magna of anesthetized rabbits, transgene activity was greater in the adventitia of intracranial arteries and meninges after injection of the complexes than adenovirus alone. Thirty minutes after application of adenovirus with the cations, binding of adenovirus to fibroblast cells in vitro or the basilar artery in vivo (by Southern blot analysis) was augmented, which suggests that enhanced binding of virus contributes to augmentation of transgene expression. Thus, cationic polymer and lipids improve transgene expression in intracranial arteries, primarily in the adventitia, after adenovirus-mediated gene transfer in vivo. This strategy may be applicable to studies of gene transfer and eventually for gene therapy.

Effects of erythrocytes and serum proteins on lung accumulation of lipoplexes containing cholesterol or DOPE as a helper lipid in the single-pass rat lung perfusion system.

Plasmid DNA-cationic liposome complexes (lipoplexes) accumulate in the lung to a great extent immediately after intravenous administration, and gene expression occurs predominantly in the lung. However, the detailed mechanisms underlying the lung accumulation of lipoplexes are not fully understood. In this study, we investigated the effect of blood components on the lung accumulation of lipoplexes using a single-pass rat lung perfusion system. Two types of lipoplexes, Chol-containing lipoplex ([(32)P]DNA-DOTMA/Chol liposome complex) and DOPE-containing lipoplex ([(32)P]DNA-DOTMA/DOPE liposome complex), pre-incubated with whole blood, serum, or erythrocytes, were injected into the perfused lung via an artery. Similarly to in vivo observations, extensive lung accumulation was observed for both types of lipoplexes after incubation with whole blood during a single passage. The (32)P-labeled lipoplexes pre-incubated with erythrocytes showed similar lung accumulation, whereas their lung accumulation after incubation with serum was significantly reduced, suggesting that erythrocytes would be more responsible blood components for extensive uptake by the perfused lung. However, there was a clear difference in the amounts of the accumulated erythrocytes after intra-arterial injection between the two lipoplex formulations. A significant degree of erythrocyte accumulation was observed when the DOPE-containing lipoplex was injected, whereas the Chol-containing lipoplex failed to induce any significant erythrocyte accumulation in the lung. In vitro experiments showed that the major fraction of both lipoplexes was bound to erythrocytes. These data suggested that Chol-containing lipoplexes bound to erythrocytes before injection dissociate from the erythrocytes and are transferred to the lung capillary endothelial cells during their passage through the lung. In contrast, DOPE-containing lipoplexes bound to erythrocytes cause aggregation and are embolized in the lung capillary with erythrocytes. Thus, the present study demonstrated that the interaction with erythrocytes plays an important role in the lung accumulation of lipoplexes and that neutral helper lipid significantly affects this interaction.

Cationic poly(ethyleneglycol) lipids incorporated into pre-formed vesicles enhance binding and uptake to BHK cells.

This paper describes a new method for enhancing the interaction of liposomes with cells. A novel class of cationic poly(ethyleneglycol) (PEG)-lipid (CPL) conjugates have been characterized for their ability to insert into pre-formed vesicles and enhance in vitro cellular binding and uptake of neutral and sterically-stabilized liposomes. The CPLs, which consist of a distearoylphosphatidylethanolamine (DSPE) anchor, a fluorescent dansyl moiety, a heterobifunctional PEG polymer (M(r) 3400), and a cationic headgroup composed of lysine derivatives, have been described previously [Bioconjug. Chem. 11 (2000) 433]. Five separate CPL, possessing 1-4 positive charges in the headgroup (referred to as CPL(1)-CPL(4), respectively), were incubated (as micellar solutions) in the presence of neutral or sterically-stabilized cationic large unilamellar vesicles (LUVs), and were found to insert into the external leaflet of the LUVs in a manner dependent on temperature, time, CPL/lipid ratio, and LUV composition. For CPL/lipid molar ratios < or =0.1, optimal insertion levels of approximately 70% of initial CPL were obtained following 3 h at 60 degrees C. The insertion of CPL resulted in aggregation of the LUVs, as assessed by fluorescence microscopy, which could be prevented by the presence of 40 mM Ca(2+). The effect of CPL-insertion on the binding of LUVs to cells was examined by fluorescence microscopy and quantified by measuring the ratio of rhodamine fluorescence to protein concentration. Neither control LUVs or LUVs containing CPL(2) displayed significant uptake by BHK cells. However, a 3-fold increase in binding was observed for LUVs possessing CPL(3), while for CPL(4)-LUVs values as high as 10-fold were achieved. Interestingly, the increase in lipid uptake did not correlate with total surface charge, but rather with increased positive charge density localized at the CPL distal headgroups. These results suggest that incorporation of CPLs into existing liposomal drug delivery systems may lead to significant improvements in intracellular delivery of therapeutic agents.

Optimization of lipid composition in cationic emulsion as in vitro and in vivo transfection agents.

PURPOSE: To enhance in vitro and in vivo transfection activity by optimizing lipid composition of cationic lipid emulsions. METHODS: Various emulsion formulations having different cationic lipids as emulsifiers, and additional helper lipids as co-emulsifiers, were prepared. The stability of the emulsion and its complex with DNA was investigated by measuring the particle size change in phosphate buffer saline (PBS) over a period of 20 days. The activity of the emulsions in transfecting pCMV-beta into COS-1 cells in the presence or absence of 80% serum was evaluated. We also evaluated in vivo transfection activity using intravenously administered pCMV-Luc+ as a reporter gene. RESULTS: Among the cationic emulsifiers, 1,2-dioleoyl-sn-glycero-3-trimethylammonium-propane (DOTAP) formed the most stable and efficient emulsion gene carrier. Addition of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) increased in vitro transfection activity, but slightly compromised the stability of the emulsion. The loss was compensated for by including small amounts of Tween 80 in the emulsion. The in vitro and in vivo transfection activities were also increased by adding Tween 80. Even though in vitro transfection activity of liposomes was high in the absence of serum, the transfection activity of emulsions was far greater than that of liposomes in the presence of serum and for in vivo applications. CONCLUSIONS: By including DOPE as an endosomolytic agent and Tween 80 as a stabilization agent, the cationic emulsion becomes a more potent gene carrier for in vitro and in vivo applications, especially in the presence of serum.