Synthesis and Characterisation of Aqueous Haemoglobin-based Microcapsules Coated by Genipin-Cross-Linked Albumin

Abstract

Figure 1. Formation of BSA/Hb-Microcapsules from co-precipitation to cross-linking and dissolution of the carrier particles.Bovine serum albumin (BSA)-coated haemoglobin (Hb)-microcapsules prepared by co-precipitation of Hb and MnCO3 may present an alternative type of artificial blood substitute. Preparation and storage in Ringer's solution provides a medium for synthesis as well as for application to the circulatory system later on. The capsules come along with a mean diameter of approximately 0.6 μm and a mean volume of 1.13 • 10 -19 L, thus an average human red blood cell with a volume of 9 • 10 -14 L is about 800,000 times bigger. Hb-microcapsules prepared by the applied method are fully regenerable by ascorbic acid and maintain oxygen affinity because oxygen is able to pass the BSA wall of the capsules and thereby binding to the ferrous iron of the haemoglobin entity. Therefore, these microcapsules present a suitable type of potential artificial haemoglobin-based oxygen carrier (HbOC).

Synthesis and characterization of enzymatically active micrometer protein-capsules

“…ACCs were created by a method first presented by Xiong et al [5,19,20] and later modified by Schakowski et al [4]. Crosslinking of the BSA molecules by genipin as described in previous works [4,[21][22][23] results in a stable shell of BSA around the core particle.…”

“…ACCs were created by a method first presented by Xiong et al [5,19,20] and later modified by Schakowski et al [4]. Crosslinking of the BSA molecules by genipin as described in previous works [4,[21][22][23] results in a stable shell of BSA around the core particle. Dissolution of MnCO 3 from the core by EDTA finally leads to the release of formerly entrapped catalase to the inside of the created capsule (Figure 1).…”

“…In order to counteract the rising shortness on blood donations all over the world, the development of artificial blood substitutes has been investigated with several different approaches [1][2][3]. Possible blood substitutes were created using either hemoglobin-based oxygen carriers (HbOCs) [4,5] or perfluorocarbon-based oxygen carriers (PFCOCs) [6]. HbOCs as well as PFCOCs come with a shell of either biodegradable polymers or albumin [5,7,8] whereas free haemoglobin has widely been ruled out due to nephrotoxicity [9].…”

“…One possible encapsulation technique for biomedical application is liposome-encapsulation of enzymes [17] but this bears the disadvantage that efficiency depends on interaction between the protein and the phospholipid bilayer [18]. The encapsulation of proteins into a shell of albumin has already successfully been performed by different research groups [5,4,19], so the encapsulation of functional enzymes should be the consistent next step in creating artificial RBCs. salt dihydrate (Na 2 EDTA) and Triton X-100 were purchased from Sigma-Aldrich (Darmstadt, Germany); Sodium carbonate anhydrous and Sodium hydroxide were purchased from Merck (Darmstadt, Germany); Disodium phosphate dehydrate was purchased from Carl Roth (Karlsruhe, Germany); Ringer's solution was purchased from Fresenius Kabi (Bad Homburg, Germany); Gibco Minimum Essential Medium (MEM), Gibco Foetal bovine serum (FBS), Gibco Pen Strep and Gibco 0.05% Trypsin-EDTA were purchased from Thermo Fisher Scientific (Schwerte, Germany); Catalase from bovine liver (ca.…”

Creation and analysis of enzymatically active micrometer protein-capsules

“…In order to counteract the rising shortness on blood donations all over the world, the development of artificial blood substitutes has been investigated with several different approaches (Müller et al 2015, Chung et al 2016, Ellingson et al 2017. Possible blood substitutes were created using either hemoglobine-based oxygen carriers (HbOCs) (Xiong et al 2012, Schakowski et al 2020 or perfluorocarbon-based oxygen carriers (PFCOCs) (Wrobeln et al 2017). HbOCs as well as PFCOCs come with a shell of either biodegradable polymers or albumin (Chang and Yu 1995, Bauer et al 2010, Xiong et al 2012 whereas free hemoglobin has widely been ruled out due to nephrotoxicity (Simoni et al 1997).…”