Nanoemulsion synthetic red blood cells having sequentially formed calcium phosphate coating and preparation method thereof

a technology of calcium phosphate coating and synthetic red blood cells, which is applied in the direction of coating, capsule delivery, microcapsules, etc., can solve the problems of increasing production costs, complex preparation process, and purified hemoglobin isolated from red blood cells

Inactive Publication Date: 2018-09-13
HAN KYU BUM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0011]It is an object of the present invention to provide nanoemulsion type synthetic red blood cells having a calcium phosphate coating of controllable thickness, in which the oxygen capacity and oxygen release rate of the synthetic red blood cells can be controlled by controlling the thickness of Ca2+ and PO42− coating layers thereon and the synthetic red blood cells can be retrieved and reused, and a preparation method thereof.

Problems solved by technology

In addition, pure hemoglobin isolated from red blood cells has a disadvantage in that oxygen is not exactly delivered.
To overcome this disadvantage, cross-linking, polymerization, encapsulation and the like should be performed during the preparation process, thus making the preparation process complex and increasing the production cost.
However, these PFC-based synthetic red blood cells have a shortcoming in that they have poor stability because they are cleared in vivo within 48 hours.
The perfluorocarbon-based synthetic red blood cells have disadvantages in that, because Ca2+ and PO42− are added at the same time in the process of coating with Ca2+ and PO42−, most of the added Ca2+ and PO42− form the ceramic crystal calcium phosphate without being attached to the submicron structures, and thus the yield of coating on the submicron structures is low, and the production process cannot always provide submicron structures having the same coating, and thus has low reproducibility.

Method used

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  • Nanoemulsion synthetic red blood cells having sequentially formed calcium phosphate coating and preparation method thereof
  • Nanoemulsion synthetic red blood cells having sequentially formed calcium phosphate coating and preparation method thereof
  • Nanoemulsion synthetic red blood cells having sequentially formed calcium phosphate coating and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

preparation example 1

Phosphate-Coated Nanoemulsion

[0060]Soy lecithin and perfluorooctyl bromide (PFOB) were respectively purchased from Avanti Polar Lipids, Inc. (Alabaster, Ala.), and Oakwood Chemical, Inc. (Alabaster, Ala.). Packed red blood cells (RBCs) were obtained from the University Hospital Blood Bank at the University of Utah. The RBCs were washed three times with normal saline before being hemolyzed with a hypotonic phosphate buffer. After centrifugation, a supernatant of hemoglobin solution was obtained.

[0061]To synthesize an oil-in-water nanoemulsion, soy lecithin (1.1 mg / g) was added to PFOB (2 vol %) and deionized water. The sample was stirred at 1200 rpm for 30 minutes at room temperature and extruded through a 200-nm pore size membrane, seven times, at 150 psi. A titration method was utilized to optimize the surface charge of the emulsion with hydrochloric acid (HCl, 0.1 M) and to observe the ionic affinity between calcium and phosphate ions; calcium chloride (CaCl2, 3.0 mM, pH 9) and ph...

example 1

on of the Characteristics and Stability of Nanoemulsion

[0062]The particle size and zeta-potential were characterized by Zetasizer Nano ZS (Malvern, UK). Transmission electron microscope (FEI Tecnai™ TEM) was utilized for morphology observations, where the emulsion was prepared with a negative stain, uranyl acetate (47 mM, 10 μL). The coated and uncoated emulsions (with and without CaP) were observed at 80 keV.

[0063]The pH titration test clearly supported the notion that the phosphate group in soy lecithin was the main site of ion interaction. Since the amine group in soy lecithin is a quaternary amine group having three methyl groups and one R group, it is not pH active. The phosphate in the head group of soy lecithin was the only group that was expected to react with a proton. The zeta-potential is a good indicator of the dispersion stability. Near zero mV results in minimal electrostatic repulsion between particles, so the greater absolute value of zeta-potential, the better the s...

example 2

on of the Rate and Amount of Oxygen Release

[0072]Packed red blood cells (RBCs) were hemolyzed with a hypotonic phosphate buffer (5 mM, pH 7.4) to obtain a hemoglobin solution whose concentration was measured by UV-Vis spectroscopy. After determining its raw concentration, the solution was adjusted to 120 μM and pH 7.2. Deoxygenated hemoglobin (Hb) was formed by purging the solution with nitrogen gas until the dissolved oxygen probe was reading zero. In order to measure the rate of oxygen release, Hb and a stopped-flow apparatus (American Instrument Co, Inc, Silver Spring, MY) with a spectrophotometer (Beckman, National Technical Laboratories) were utilized. Four different samples were produced, each with a different number of calcium and phosphate layers. The apparatus was a light-beam path of 4 mm and the slit (0.82 mm). The transmittance of the mixture was recorded using LabView software. By monitoring the change of transmittance at 550 nm, it was shown that Hb was changed to oxyg...

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Abstract

The present invention relates to nanoemulsion type synthetic red blood cells having a calcium phosphate coating of controllable thickness, and to a preparation method thereof. According to the present invention, Ca2+ and PO42− coating layers are uniformly and sequentially formed using a layer-by-layer (LBL) coating method while controlling the thicknesses of the coating layers. Thus, the oxygen capacity and oxygen release rate of the synthetic red blood cells can be controlled, and the synthetic red blood cells can be retrieved and reused.

Description

CROSS-REFERENCE TO RELATED APPLICATION[0001]This application claims priority under 35 U.S.C. § 119 of Korean Patent Application No. 10-2017-0030524 filed Mar. 10, 2017. The disclosure of such Korean priority patent application is hereby incorporated herein by reference in its entirety, for all purposes.TECHNICAL FIELD[0002]The present invention relates to nanoemulsion type synthetic red blood cells having a calcium phosphate coating formed by sequential coating, and a preparation method thereof. More specifically, the present invention relates to nanoemulsion type synthetic red blood cells having a calcium phosphate coating formed by sequential coating, in which a Ca2+ coating layer and a PO42− coating layer are uniformly and sequentially formed as separate layers by use of a layer-by-layer (LBL) coating method while controlling the thicknesses of the calcium and phosphate coating layers so that the oxygen capacity and oxygen release rate of the synthetic red blood cells can be cont...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61K31/685A61K9/107A61K9/00A61K9/14B05D1/26B05D3/00
CPCA61K31/685A61K9/107A61K9/0026B05D3/007A61K9/145B05D1/265A61K9/143A61K9/06A61K9/1075A61K9/501A61K47/06A61K47/24
Inventor HAN, KYU-BUM
Owner HAN KYU BUM
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