Semi-Autonomic AI LF-NMR Sensor for Industrial Prediction of Edible Oil Oxidation Status.

The evaluation of an oil's oxidation status during industrial production is highly important for monitoring the oil's purity and nutritional value during production, transportation, storage, and cooking. The oil and food industry is seeking a real-time, non-destructive, rapid, robust, and low-cost sensor for nutritional oil's material characterization. Towards this goal, a 1H LF-NMR relaxation sensor application based on the chemical and structural profiling of non-oxidized and oxidized oils was developed. This study dealt with a relatively large-scale oil oxidation database, which included crude data of a 1H LF-NMR relaxation curve, and its reconstruction into T1 and T2 spectral fingerprints, self-diffusion coefficient D, and conventional standard chemical test results. This study used a convolutional neural network (CNN) that was trained to classify T2 relaxation curves into three ordinal classes representing three different oil oxidation levels (non-oxidized, partial oxidation, and high level of oxidation). Supervised learning was used on the T2 signals paired with the ground-truth labels of oxidation values as per conventional chemical lab oxidation tests. The test data results (not used for training) show a high classification accuracy (95%). The proposed AI method integrates a large training set, an LF-NMR sensor, and a machine learning program that meets the requirements of the oil and food industry and can be further developed for other applications.

1H LF-NMR Self-Diffusion Measurements for Rapid Monitoring of an Edible Oil's Food Quality with Respect to Its Oxidation Status.

The food quality of edible oils is dependent on basic chemical and structural changes that can occur by oxidation during preparation and storage. A rapid and efficient analytical method of the different steps of oil oxidation is described using a time-domain nuclear magnetic resonance (TD-NMR) sensor for measuring signals related to the chemical and physical properties of the oil. The degree of thermal oxidation of edible oils at 80 °C was measured by the conventional methodologies of peroxide and aldehyde analysis. Intact non-modified samples of the same oils were more rapidly analyzed for oxidation using a TD-NMR sensor for 2D T1-T2 and self-diffusion (D) measurements. A good linear correlation between the D values and the conventional chemical analysis was achieved, with the highest correlation of R2 = 0.8536 for the D vs. the aldehyde concentrations during the thermal oxidation of poly-unsaturated linseed oils, the oil most susceptible to oxidation. A good correlation between the D and aldehyde levels was also achieved for all the other oils. The possibility to simplify and minimize the time of oxidative analysis using the TD NMR sensors D values is discussed as an indicator of the oil's oxidation quality, as a rapid and accurate methodology for the oil industry.

Time Domain (TD) Proton NMR Analysis of the Oxidative Safety and Quality of Lipid-Rich Foods.

Food safety monitoring is highly important due to the generation of unhealthy components within many food products during harvesting, processing, storage, transportation and cooking. Current technologies for food safety analysis often require sample extraction and the modification of the complex chemical and morphological structures of foods, and are either time consuming, have insufficient component resolution or require costly and complex instrumentation. In addition to the detection of unhealthy chemical toxins and microbes, food safety needs further developments in (a) monitoring the optimal nutritional compositions in many different food categories and (b) minimizing the potential chemical changes of food components into unhealthy products at different stages from food production until digestion. Here, we review an efficient methodology for overcoming the present analytical limitations of monitoring a food's composition, with an emphasis on oxidized food components, such as polyunsaturated fatty acids, in complex structures, including food emulsions, using compact instruments for simple real-time analysis. An intelligent low-field proton NMR as a time domain (TD) NMR relaxation sensor technology for the monitoring of T2 (spin-spin) and T1 (spin-lattice) energy relaxation times is reviewed to support decision-making by producers, retailers and consumers in regard to food safety and nutritional value during production, shipping, storage and consumption.

Proton Low Field NMR Relaxation Time Domain Sensor for Monitoring of Oxidation Stability of PUFA-Rich Oils and Emulsion Products.

The nutritional characteristics of fatty acid (FA) containing foods are strongly dependent on the FA's chemical/morphological arrangements. Paradoxically the nutritional, health enhancing FA polyunsaturated fatty acids (PUFAs) are highly susceptible to oxidation into harmful toxic side products during food preparation and storage. Current analytical technologies are not effective in the facile characterization of both the morphological and chemical structures of PUFA domains within materials for monitoring the parameters affecting their oxidation and antioxidant efficacy. The present paper is a review of our work on the development and application of a proton low field NMR relaxation sensor (1H LF NMR) and signal to time domain (TD) spectra reconstruction for chemical and morphological characterization of PUFA-rich oils and their oil in water emulsions, for assessing their degree and susceptibility to oxidation and the efficacy of antioxidants. The NMR signals are energy relaxation signals generated by spin-lattice interactions (T1) and spin-spin interactions (T2). These signals are reconstructed into 1D (T1 or T2) and 2D graphics (T1 vs. T2) by an optimal primal-dual interior method using a convex objectives (PDCO) solver. This is a direct measurement on non-modified samples where the individual graph peaks correlate to structural domains within the bulk oil or its emulsions. The emulsions of this review include relatively complex PUFA-rich oleosome-oil bodies based on the aqueous extraction from linseed seeds with and without encapsulation of externally added oils such as fish oil. Potential applications are shown in identifying optimal health enhancing PUFA-rich food formulations with maximal stability against oxidation and the potential for on-line quality control during preparation and storage.

Cationic vesicles from novel bolaamphiphilic compounds.

Effective targeted drug delivery by cationic liposomes is difficult to achieve because of their rapid clearance from the blood circulation. Bolaamphiphiles that form monolayer membrane may provide vesicles with improved stability, as shown for archaeosomes. We investigated a series of bolaamphiphiles with acetylcholine head groups and systematic structural changes in their hydrophobic domain for their ability to form stable nanovesicles. Bolaamphiphiles with two aliphatic chains separated by a short amide midsection produced spherical nanovesicles ranging in diameter from 80 to 120 nm. These vesicles lost their encapsulated material within 24 hours of incubation in phosphate-buffered saline (PBS). Similar bolaamphiphiles with a longer midsection produced a mixture of fibers and more stable nanovesicles. Bolaamphiphiles with ester amide midsection produced only spherical nanovesicles that were stable during incubation in PBS for several days. Vesicles made from bolaamphiphiles with acetylcholine head groups conjugated to the aliphatic chain via the amine were less stable than vesicles made from bolaamphiphiles with head groups conjugated to the aliphatic chain via the acetyl group. Vesicles that were stable in vitro showed good stability in the blood circulation after intravenous administration to mice. These results help in elucidating the bolaamphiphile structures needed to form stable cationic vesicles for targeted drug delivery.

Characterization of Cationic Bolaamphiphile Vesicles for siRNA Delivery into Tumors and Brain.

Small interfering RNAs (siRNAs) are potential therapeutic substances due to their gene silencing capability as exemplified by the recent approval by the US Food and Drug Administration (FDA) of the first siRNA therapeutic agent (patisiran). However, the delivery of naked siRNAs is challenging because of their short plasma half-lives and poor cell penetrability. In this study, we used vesicles made from bolaamphiphiles (bolas), GLH-19 and GLH-20, to investigate their ability to protect siRNA from degradation by nucleases while delivering it to target cells, including cells in the brain. Based on computational and experimental studies, we found that GLH-19 vesicles have better delivery characteristics than do GLH-20 vesicles in terms of stability, binding affinity, protection against nucleases, and transfection efficiency, while GLH-20 vesicles contribute to efficient release of the delivered siRNAs, which become available for silencing. Our studies with vesicles made from a mixture of the two bolas (GLH-19 and GLH-20) show that they were able to deliver siRNAs into cultured cancer cells, into a flank tumor and into the brain. The vesicles penetrate cell membranes and the blood-brain barrier (BBB) by endocytosis and transcytosis, respectively, mainly through the caveolae-dependent pathway. These results suggest that GLH-19 strengthens vesicle stability, provides protection against nucleases, and enhances transfection efficiency, while GLH-20 makes the siRNA available for gene silencing.

Novel cationic vesicle platform derived from vernonia oil for efficient delivery of DNA through plant cuticle membranes.

Novel cationic amphiphilic compounds were prepared from vernonia oil, a natural epoxidized triglyceride, and studied with respect to vesicle formation, encapsulation of biomaterials such as DNA, and their physical stability and transport through isolated plant cuticle membranes. The amphiphiles studied were a single-headed compound III (a quaternary ammonium head group with two alkyl chains) and a triple-headed compound IV, which is essentially three molecules of compound III bound together through a glycerol moiety. Vesicles of the two amphiphiles, prepared by sonication in water and solutions of uranyl acetate or the herbicide 2,4-D (2,4-dichloropenoxy acetic acid), were examined by TEM, SEM, AFM, and confocal laser systems and had a spherical shape which encapsulated the solutes with diameters between 40 and 110 nm. Vesicles from amphiphile IV could be made large enough to encapsulate a condensed 5.2kb DNA plasmid (pJD328). Vesicles of amphiphile IV were also shown to pass intact across isolated plant cuticle membranes and the rate of delivery of encapsulated radio-labeled 2,4-D through isolated plant cuticle membranes obtained with these vesicles was clearly greater in comparison to liposomes prepared from dipalmitopyl phosphatidylcholine (DPPC) and the control, nonencapsulated 2,4-D. Vesicles from amphiphiles III and IV were found to be more stable than those of liposomes from DPPC. The data indicate the potential of vesicles prepared from the novel amphiphile IV to be a relatively efficient nano-scale delivery system to transport DNA and other bioactive agents through plant biological barriers. This scientific approach may open the way for further development of efficient in vivo plant transformation systems.

Liquid-phase characterization of molecular interactions in polyunsaturated and n-fatty acid methyl esters by (1)H low-field nuclear magnetic resonance.

BACKGROUND: To identify and develop the best renewable and low carbon footprint biodiesel substitutes for petroleum diesel, the properties of different biodiesel candidates should be studied and characterized with respect to molecular structures versus biodiesel liquid property relationships. In our previous paper, (1)H low-field nuclear magnetic resonance (LF-NMR) relaxometry was investigated as a tool for studying the liquid-phase molecular packing interactions and morphology of fatty acid methyl esters (FAMEs). The technological potential was demonstrated with oleic acid and methyl oleate standards having similar alkyl chains but different head groups. In the present work, molecular organization versus segmental and translational movements of FAMEs in their pure liquid phase, with different alkyl chain lengths (10-20 carbons) and degrees of unsaturation (0-3 double bonds), were studied with (1)H LF-NMR relaxometry and X-ray, (1)H LF-NMR diffusiometry, and (13)C high-field NMR. RESULTS: Based on density values and X-ray measurements, it was proposed that FAMEs possess a liquid crystal-like order above their melting point, consisting of random liquid crystal aggregates with void spaces between them, whose morphological properties depend on chain length and degree of unsaturation. FAMEs were also found to exhibit different degrees of rotational and translational motions, which were rationalized by chain organization within the clusters, and the degree and type of molecular interactions and temperature effects. At equivalent fixed temperature differences from melting point, saturated FAME molecules were found to have similar translational motion regardless of chain length, expressed by viscosity, self-diffusion coefficients, and spin-spin (T 2) (1)H LF-NMR. T 2 distributions suggest increased alkyl chain rigidity, and reduced temperature response of the peaks' relative contribution with increasing unsaturation is a direct result of the alkyl chain's morphological packing and molecular interactions. CONCLUSIONS: Both the peaks' assignments for T 2 distributions of FAMEs and the model for their liquid crystal-like morphology in the liquid phase were confirmed. The study of morphological structures within liquids and their response to temperature changes by (1)H LF-NMR has a high value in the field of biodiesel and other research and applied disciplines in numerous physicochemical- and organizational-based properties, processes, and mechanisms of alkyl chains, molecular interactions, and morphologies.

Study of liquid-phase molecular packing interactions and morphology of fatty acid methyl esters (biodiesel).

BACKGROUND: (1)H low field nuclear magnetic resonance (LF-NMR) relaxometry has been suggested as a tool to distinguish between different molecular ensembles in complex systems with differential segmental or whole molecular motion and/or different morphologies. In biodiesel applications the molecular structure versus liquid-phase packing morphologies of fatty acid methyl esters (FAMEs) influences physico-chemical characteristics of the fuel, including flow properties, operability during cold weather, blending, and more. Still, their liquid morphological structures have scarcely been studied. It was therefore the objective of this work to explore the potential of this technology for characterizing the molecular organization of FAMEs in the liquid phase. This was accomplished by using a combination of supporting advanced technologies. RESULTS: We show that pure oleic acid (OA) and methyl oleate (MO) standards exhibited both similarities and differences in the (1)H LF-NMR relaxation times (T2s) and peak areas, for a range of temperatures. Based on X-ray measurements, both molecules were found to possess a liquid crystal-like order, although a larger fluidity was found for MO, because as the temperature is increased, MO molecules separate both longitudinally and transversely from one another. In addition, both molecules exhibited a preferred direction of diffusion based on the apparent hydrodynamic radius. The close molecular packing arrangement and interactions were found to affect the translational and segmental motions of the molecules, as a result of dimerization of the head group in OA as opposed to weaker polar interactions in MO. CONCLUSIONS: A comprehensive model for the liquid crystal-like arrangement of FAMEs in the liquid phase is suggested. The differences in translational and segmental motions of the molecules were rationalized by the differences in the (1)H LF-NMR T2 distributions of OA and MO, which was further supported by (13)C high field (HF)-NMR spectra and (1)H HF-NMR relaxation. The proposed assignment allows for material characterization based on parameters that contribute to properties in applications such as biodiesel fuels.

Bolaamphiphiles as carriers for siRNA delivery: From chemical syntheses to practical applications.

In this study we have investigated a new class of cationic lipids--"bolaamphiphiles" or "bolas"--for their ability to efficiently deliver small interfering RNAs (siRNAs) to cancer cells. The bolas of this study consist of a hydrophobic chain with one or more positively charged head groups at each end. Recently, we reported that micelles of the bolas GLH-19 and GLH-20 (derived from vernonia oil) efficiently deliver siRNAs, while having relatively low toxicities in vitro and in vivo. Our previous studies validated that; bolaamphiphiles can be designed to vary the magnitude of siRNA shielding, its delivery, and its subsequent release. To further understand the structural features of bolas critical for siRNAs delivery, new structurally related bolas (GLH-58 and GLH-60) were designed and synthesized from jojoba oil. Both bolas have similar hydrophobic domains and contain either one, in GLH-58, or two, in GLH-60 positively charged head groups at each end of the hydrophobic core. We have computationally predicted and experimentally validated that GLH-58 formed more stable nano sized micelles than GLH-60 and performed significantly better in comparison to GLH-60 for siRNA delivery. GLH-58/siRNA complexes demonstrated better efficiency in silencing the expression of the GFP gene in human breast cancer cells at concentrations of 5µg/mL, well below the toxic dose. Moreover, delivery of multiple different siRNAs targeting the HIV genome demonstrated further inhibition of virus production.

Progress in lipid-based nanoparticles for cancer therapy.

Most conventional cancer therapeutics gain limited access to many types of tumors while having considerable adverse effects, resulting in low therapeutic efficacy and high toxicity. Therefore, research has now focused on the development of novel drug delivery systems (DDS) with the goal of maintaining high therapeutic drug levels at malignant cells and as low as possible drug levels in other cells. The introduction of nanotechnology has addressed some of these problems and opened up new avenues for improved cancer therapy. The design of nanoparticles for DDS takes into consideration issues such as targeting, controlled drug release and enhanced penetration via biological barriers. In this review we describe the design principles of targeted DDS for cancer therapy and the types of nanoparticles that are under development. Emphasis is put on lipid-based nanoparticles, particularly bolaamphiphilic vesicles that have tremendous potential in delivering therapeutic and diagnostic agents to specific cells following systemic administration.

Steric environment around acetylcholine head groups of bolaamphiphilic nanovesicles influences the release rate of encapsulated compounds.

Two bolaamphiphilic compounds with identical acetylcholine (ACh) head groups, but with different lengths of an alkyl chain pendant adjacent to the head group, as well as differences between their hydrophobic skeleton, were investigated for their ability to self-assemble into vesicles that release their encapsulated content upon hydrolysis of their head groups by acetylcholinesterase (AChE). One of these bolaamphiphiles, synthesized from vernolic acid, has an alkyl chain pendant of five methylene groups, while the other, synthesized from oleic acid, has an alkyl chain pendant of eight methylene groups. Both bolaamphiphiles formed stable spherical vesicles with a diameter of about 130 nm. The ACh head groups of both bolaamphiphiles were hydrolyzed by AChE, but the hydrolysis rate was significantly faster for the bolaamphiphile with the shorter aliphatic chain pendant. Likewise, upon exposure to AChE, vesicles made from the bolaamphiphile with the shorter alkyl chain pendant released their encapsulated content faster than vesicles made from the bolaamphiphile with the longer alkyl chain pendant. Our results suggest that the steric environment around the ACh head group of bolaamphiphiles is a major factor affecting the hydrolysis rate of the head groups by AChE. Attaching an alkyl chain to the bolaamphiphile near the ACh head group allows self-assembled vesicles to form with a controlled release rate of the encapsulated materials, whereas shorter alkyl chains enable a faster head group hydrolysis, and consequently faster release, than longer alkyl chains. This principle may be implemented in the design of bolaamphiphiles for the formation of vesicles for drug delivery with desired controlled release rates.

Delivery of analgesic peptides to the brain by nano-sized bolaamphiphilic vesicles made of monolayer membranes.

Inefficient drug delivery to the brain is a major obstacle for pharmacological management of brain diseases. We investigated the ability of bolavesicles - monolayer membrane vesicles self-assembled from synthetic bolaamphiphiles that contain two hydrophilic head groups at each end of a hydrophobic alkyl chain - to permeate the blood-brain barrier and to deliver the encapsulated materials into the brain. Cationic vesicles with encapsulated kyotorphin and leu-enkephalin (analgesic peptides) were prepared from the bolalipids GLH-19 and GLH-20 and studied for their analgesic effects in vivo in experimental mice. The objectives were to determine: (a) whether bolavesicles can efficiently encapsulate analgesic peptides, (b) whether bolavesicles can deliver these peptides to the brain in quantities sufficient for substantial analgesic effect, and to identify the bolavesicle formulation/s that provides the highest analgetic efficiency. The results indicate that the investigated bolavesicles can deliver analgesic peptides across the blood-brain barrier and release them in the brain in quantities sufficient to elicit efficient and prolonged analgesic activity. The analgesic effect is enhanced by using bolavesicles made from a mixture the bolas GLH-19 (that contains non-hydrolyzable acetylcholine head group) and GLH-20 (that contains hydrolysable acetylcholine head group) and by incorporating chitosan pendants into the formulation. The release of the encapsulated materials (the analgesic peptides kyotorphin and leu-enkephalin) appears to be dependent on the choline esterase (ChE) activity in the brain vs. other organs and tissues. Pretreatment of experimental animals with pyridostigmine (the BBB-impermeable ChE inhibitor) enhances the analgesic effects of the studied formulations. The developed formulations and the approach for their controlled decapsulation can serve as a useful modality for brain delivery of therapeutically-active compounds.

Bolaamphiphilic vesicles encapsulating iron oxide nanoparticles: new vehicles for magnetically targeted drug delivery.

Bolaamphiphiles - amphiphilic molecules consisting of two hydrophilic headgroups linked by a hydrophobic chain - form highly stable vesicles consisting of a monolayer membrane that can be used as vehicles to deliver drugs across biological membranes, particularly the blood-brain barrier (BBB). We prepared new vesicles comprising bolaamphiphiles (bolavesicles) that encapsulate iron oxide nanoparticles (IONPs) and investigated their suitability for targeted drug delivery. Bolavesicles displaying different headgroups were studied, and the effect of IONP encapsulation upon membrane interactions and cell uptake were examined. Experiments revealed more pronounced membrane interactions of the bolavesicles assembled with IONPs. Furthermore, enhanced internalization and stability of the IONP-bolavesicles were observed in b.End3 brain microvessel endothelial cells - an in vitro model of the blood-brain barrier. Our findings indicate that embedded IONPs modulate bolavesicles' physicochemical properties, endow higher vesicle stability, and enhance their membrane permeability and cellular uptake. IONP-bolavesicles thus constitute a promising drug delivery platform, potentially targeted to the desired location using external magnetic field.

In Silico, In Vitro, and In Vivo Studies Indicate the Potential Use of Bolaamphiphiles for Therapeutic siRNAs Delivery.

Specific small interfering RNAs (siRNAs) designed to silence different oncogenic pathways can be used for cancer therapy. However, non-modified naked siRNAs have short half-lives in blood serum and encounter difficulties in crossing biological membranes due to their negative charge. These obstacles can be overcome by using siRNAs complexed with bolaamphiphiles, consisting of two positively charged head groups that flank an internal hydrophobic chain. Bolaamphiphiles have relatively low toxicities, long persistence in the blood stream, and most importantly, in aqueous conditions can form poly-cationic micelles thus, becoming amenable to association with siRNAs. Herein, two different bolaamphiphiles with acetylcholine head groups attached to an alkyl chain in two distinct configurations are compared for their abilities to complex with siRNAs and deliver them into cells inducing gene silencing. Our explicit solvent molecular dynamics (MD) simulations showed that bolaamphiphiles associate with siRNAs due to electrostatic, hydrogen bonding, and hydrophobic interactions. These in silico studies are supported by various in vitro and in cell culture experimental techniques as well as by some in vivo studies. Results demonstrate that depending on the application, the extent of siRNA chemical protection, delivery efficiency, and further intracellular release can be varied by simply changing the type of bolaamphiphile used.Molecular Therapy-Nucleic Acids (2013) 2, e80; doi:10.1038/mtna.2013.5; published online 19 March 2013.

Interfacial and self-assembly properties of bolaamphiphilic compounds derived from a multifunctional oil.

The self-assembly characteristics in aqueous solutions of cationic bolaamphiphiles with systematic changes in their chemical structure is described with respect to their interfacial properties within water and at the air/water interface. Six cationic bolaamphiphiles were synthesized from multifunctional vernonia oil with the following variations: (a) two different alkyl chain lengths connecting the head groups, (b) polar ester or hydrogen bonding amide groups within the hydrophobic domain, and (c) an acetylcholine cationic head group with different conjugation sites to the alkyl chain. Surface tension measurements were used for determining critical aggregation concentration (CAC) values and air/water interfacial parameters such as 'effectiveness', surface excess concentration and area occupied by one molecule in the air/water interface. Fluorescent studies with pyrene were used to characterize CAC properties within the aqueous volume and transmission electron microscopy (TEM) for determining the aggregate structure's size, homogeneity and morphology. A bolaamphiphile molecular structure vs. interfacial property relationship was derived from this data which could be used to determine the molecular structure properties needed to generate interfacial forces to form either spherical vesicles or fibrous networks. The effects of the aliphatic chain length, head group orientation and functional groups within the hydrophobic domain on CAC, surface tension properties and self-aggregate morphology are described. Most bolaamphiphiles studied had CAC values in the 10-190 µM range, while two out of the six were found to assemble into MLM spherical vesicles with diameters ranging up to 120 nm suitable for drug delivery applications. Others formed a gelatinous network of fibers or multi-lamellar vesicles.

Delivery of proteins to the brain by bolaamphiphilic nano-sized vesicles.

Bolaamphiphilic cationic vesicles with acetylcholine (ACh) surface groups were investigated for their ability to deliver a model protein-bovine serum albumin conjugated to fluorescein isothiocyanate (BSA-FITC) across biological barriers in vitro and in vivo. BSA-FITC-loaded vesicles were internalized into cells in culture, including brain endothelial b.End3 cells, at 37 °C, but not at 4 °C, indicating an active uptake process. To examine if BSA-FITC-loaded vesicles were stable enough for in vivo delivery, we tested vesicle stability in whole serum. The half-life of cationic BSA-FITC-loaded vesicles with ACh surface groups that are hydrolyzed by choline esterase (ChE) was about 2 h, whereas the half-life of vesicles with similar surface groups, but which are not hydrolyzed by choline esterase (ChE), was over 5 h. Pyridostigmine, a choline esterase inhibitor that does not penetrate the blood-brain barrier (BBB), increased the stability of the ChE-sensitive vesicles to 6 h but did not affect the stability of vesicles with ACh surface groups that are not hydrolyzed by ChE. Following intravenous administration to pyridostigmine-pretreated mice, BSA-FITC encapsulated in ChE-sensitive vesicles was distributed into various tissues with marked accumulation in the brain, whereas non-encapsulated (free) BSA-FITC was detected only in peripheral tissues, but not in the brain. These results show that cationic bolaamphiphilic vesicles with ACh head groups are capable of delivering proteins across biological barriers, such as the cell membrane and the blood-brain barrier (BBB). Brain ChE activity destabilizes the vesicles and releases the encapsulated protein, enabling its accumulation in the brain.

Site-directed decapsulation of bolaamphiphilic vesicles with enzymatic cleavable surface groups.

Stable nano-sized vesicles with a monolayer encapsulating membrane were prepared from novel bolaamphiphiles with choline ester head groups. The head groups were covalently bound to the alkyl chain of the bolaamphiphiles either via the nitrogen atom of the choline moiety, or via the choline ester's methyl group. Both types of bolaamphiphiles competed with acetylthiocholine for binding to acetylcholine esterase (AChE), yet, only the choline ester head groups bound to the alkyl chain via the nitrogen atom of the choline moiety were hydrolyzed by the enzyme. Likewise, only vesicles composed of bolaamphiphiles with head groups that were hydrolyzed by AChE released their encapsulated material upon exposure to the enzyme. Injection of carboxyfluorescein (CF)-loaded vesicles with cleavable choline ester head groups into mice resulted in the accumulation of CF in tissues that express high AChE activity, including the brain. By comparison, when vesicles with choline ester head groups that are not hydrolyzed by AChE were injected into mice, there was no accumulation of CF in tissues that highly express the enzyme. These results imply that bolaamphiphilic vesicles with surface groups that are substrates to enzymes which are highly expressed in target organs may potentially be used as a drug delivery system with controlled site-directed drug release.