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The search for artificial blood substitutes that are suitable for safe transfusion in clinical conditions and in extreme situations has gained increasing interest during recent years. Most of the problems related to donor blood could be overcome with hemoglobin sub-micron particles (HbMPs) that are able to bind and deliver oxygen. On the other hand, the length of the circulation time of HbMPs in the bloodstream strongly depends on their surface properties and can be improved with biopolymer coatings. The redox potential of HbMPs and HbMPs coated with biopolymers using the layer-by-layer technique (LbL-HbMPs) is related to the energy required for electron transfer upon transition from an oxidized to a reduced state. It can be used as a measure of the stability of Hb against oxidation, which is directly connected with its function as an oxygen carrier. The redox potential of Hb, HbMPs, and LbL-HbMPs was determined by a spectroelectrochemical method utilizing the shift of the Soret peak of Hb upon oxidation/reduction of the iron in the heme. The obtained results showed a slight shift in the redox potential of both particle types of about 17 mV towards more negative values compared to the free Hb in the solution. It was demonstrated that the free Hb and the cross-linked Hb in HbMPs and LbL-HbMPs undergo transitions from an oxidized to a reduced state and vice versa several times without Hb destruction. The LbL coating does not affect the redox properties of HbMPs. This ability, as well as the proximity of the obtained redox potentials of Hb, HbMPs, and LbL-HbMPs, indicates that the eventual oxidation of HbMPs in the bloodstream is reversible; thus, HbMPs can be active as artificial oxygen carriers for a longer period of time.

Hemoglobin-based oxygen carriers (HBOCs) as blood substitutes are one of the great hopes of modern transfusion and emergency medicine. After the major safety-relevant challenges of the last decades seem to be largely overcome, current developments have in common that they are affected by degradation and excretion at an early stage in test organisms. Several possible mechanisms that may be responsible for this are discussed in the literature. One of them is CD163, the receptor of the complex of haptoglobin (Hp) and hemoglobin (Hb). The receptor has been shown in various studies to have a direct affinity for Hb in the absence of Hp. Thus, it seems reasonable that CD163 could possibly also bind Hb within HBOCs and cause phagocytosis of the particles. In this work we investigated the role of CD163 in the uptake of our hemoglobin sub-micron particles (HbMPs) in monocytes and additionally screened for alternative ways of particle recognition by monocytes. In our experiments, blockade of CD163 by specific monoclonal antibodies proved to partly inhibit HbMP uptake by monocytes. It appears, however, that several other phagocytosis pathways for HbMPs might exist, independent of CD163 and also Hb.

Doxorubicin (DOX) is an effective anthracycline antibiotic drug which is commonly used in a broad range cancer therapy. However, due to dose depending side effects and toxicity to non-cancerous tissues, its clinical applications are restricted. To overcome these limitations, human serum albumin (HSA) has been investigated as a biocompatible drug delivery vehicle. In this study, human serum albumin submicron particles (HSA-MPs) were fabricated by using the Co-precipitation–Crosslinking–Dissolution technique (CCD technique) and DOX was loaded into the protein particles by absorption. DOX-HSA-MPs showed uniform peanut-like shape, submicron size and negative zeta-potential (−13 mV). The DOX entrapment efficiency was 25% of the initial amount. The in vitro release in phosphate buffered saline pH 7.4 was less than 1% within 5 h. In contrast, up to 40% of the entrapped DOX was released in presence of a protein digesting enzyme mixture (Pronase®) within the same time. In addition, in vitro cytotoxicity and cellular uptake of DOX-HSA-MPs were evaluated using the lung carcinoma cell line A549. The results demonstrated that DOX-HSA-MPs reduced the cell metabolic activities after 72 h. Interestingly, DOX-HSA-MPs were taken up by A549 cells up to 98% and localized in the cell lysosomal compartment. This study suggests that DOX-HSA-MPs which was fabricated by CCD technique is seen as a promising biopolymer particle as well as a viable alternative for drug delivery application to use for cancer therapy.

Superoxide dismutase (SOD) and Catalase (CAT) play a crucial role as the first line of defense antioxidant enzymes in a living cell. These enzymes neutralize the superoxide anion from the autooxidation of oxyhemoglobin (Oxy-Hb) and convert hydrogen peroxides into water and molecular oxygen. In this study, we fabricated hemoglobin submicron particles (HbMPs) using the Coprecipitation Crosslinking Dissolution (CCD) technique and incorporating first-line antioxidant enzymes (CAT, SOD) and second-line antioxidant (ascorbic acid, Vit. C) to investigate a protective effect of modified HbMPs via cyclically oxygenation and deoxygenation. Thereafter, the total hemoglobin (Hb) content and Oxy-Hb content to HbMPs were determined. The results revealed that the HbMPs have a protective effect against oxidation from hydrogen peroxide and potentially neutralizing hydrogen peroxide to water over 16 times exposure cycles. No significant differences in total Hb content were found between normal HbMPs and enzyme-modified HbMPs in the absence of Vit. C. The Oxy-Hb of CAT-HbMPs showed significantly higher values than normal HbMPs. The functional Hb of normal HbMPs and enzyme-modified HbMPs was increased by 60–77% after a short time Vit. C (1:25) exposure. The co-immobilization of CAT and SOD in hemoglobin particles (CAT-SOD-HbMPs) in the presence of Vit. C provides protective effects against oxidation in cyclic Oxygenation and Deoxygenation and shows the lowest reduction of functional Hb. Our studies show that the CCD technique-modified HbMPs containing antioxidant enzymes and a reducing agent (ascorbic acid) demonstrate enhanced Hb functionality, providing protective effects and stability under oxidative conditions.

The use of nanoparticles is becoming increasingly apparent in a growing number of medical fields. To exploit the full potential of these particles, it is essential to examine their behavior in the blood and their possible interactions with blood cells. Dendritic core multi-shell DendroSol™ nanoparticles (DS-NPs) are able to penetrate into viable layers of human skin, but nothing is known about their interaction with blood cells. In the present study, we analyze the effect of DS-NPs on red blood cells (RBCs) using confocal laser scanning microscopy (CLSM), flow cytometry, sedimentation rate analysis, spectrophotometry, and hemolysis assays. DS-NPs labeled with Nile red (NR) were added to RBC suspensions and their accumulation in the area of the RBC membranes was demonstrated by CLSM as well as by flow cytometry. In the presence of DS-NPs, the RBCs show an increased sedimentation rate, which also confirms the binding of the NPs to the cells. Interestingly, in the presence of DS-NPs, the RBCs are stabilized against hypotonic hemolysis as well as against the hemolytic action of Triton X-100. This proven anti-hemolytic effect could be utilized to enhance the circulation time of RBCs loaded with drugs for prolonged sustained release and drug delivery with enhanced bioavailability.

Although riboflavin (RF) belongs to the water-soluble vitamins of group B, its solubility is low. Therefore, the application of micro-formulations may help to overcome this limiting factor for the delivery of RF. In this study we immobilized RF in newly developed albumin submicron particles prepared using the Co-precipitation Crosslinking Dissolution technique (CCD-technique) of manganese chloride and sodium carbonate in the presence of human serum albumin (HSA) and RF. The resulting RF containing HSA particles (RF-HSA-MPs) showed a narrow size distribution in the range of 0.9 to 1 μm, uniform peanut-like morphology, and a zeta-potential of −15 mV. In vitro release studies represented biphasic release profiles of RF in a phosphate buffered saline (PBS) pH 7.4 and a cell culture medium (RPMI) 1640 medium over a prolonged period. Hemolysis, platelet activation, and phagocytosis assays revealed a good hemocompatibility of RF-HSA-MPs.

In this research, we aimed to assess antibacterial activity and develop oral care products from three natural plant extracts from the Thai highlands. The plants, including Camellia sinensis var. assamica, Zanthozylum limonella Alston, and Acorus calamus L., were extracted using two traditional extraction techniques: maceration and hydrodistillation methods. The extracts were characterized by percentage yield, total phenolic, and total flavonoid contents. Antibacterial activity against Staphylococcus aureus, which play a role in oral health and disease, was investigated. C. sinensis var. assamica extract had the highest content of phenolic acid (38.15 ± 4.12 mg GAE/g extract) and flavonoids (44.91 ± 2.76 mg QE/g extract). Interestingly, a combination of C. sinensis with Z. limonella and A. calamus provides a greater inhibitory effect against S. aureus. Furthermore, oral care products were prepared as a natural product mixture in two preparations: (i) oral ulcers gel and (ii) oral spray. Apart from antibacterial efficiency, volunteer satisfaction after the usage of oral care products containing traditional plant extracts was investigated via organoleptic evaluation. The findings of the volunteer surveys indicated positive feedback for both oral care products with high satisfaction levels. Hence, these oral care products could potentially be natural antimicrobial agents and can be further developed and applied for oral applications in the pharmaceutical and cosmetic industries.

In this research, we aimed to assess antibacterial activity and develop oral care products from three natural plant extracts from the Thai highlands. The plants, including Camellia sinensis var. assamica, Zanthozylum limonella Alston, and Acorus calamus L., were extracted using two traditional extraction techniques: maceration and hydrodistillation methods. The extracts were characterized by percentage yield, total phenolic, and total flavonoid contents. Antibacterial activity against Staphylococcus aureus, which play a role in oral health and disease, was investigated. C. sinensis var. assamica extract had the highest content of phenolic acid (38.15 ± 4.12 mg GAE/g extract) and flavonoids (44.91 ± 2.76 mg QE/g extract). Interestingly, a combination of C. sinensis with Z. limonella and A. calamus provides a greater inhibitory effect against S. aureus. Furthermore, oral care products were prepared as a natural product mixture in two preparations: (i) oral ulcers gel and (ii) oral spray. Apart from antibacterial efficiency, volunteer satisfaction after the usage of oral care products containing traditional plant extracts was investigated via organoleptic evaluation. The findings of the volunteer surveys indicated positive feedback for both oral care products with high satisfaction levels. Hence, these oral care products could potentially be natural antimicrobial agents and can be further developed and applied for oral applications in the pharmaceutical and cosmetic industries.

Non-sericin (NS) extract was produced from the ethanolic extract of Bombyx mori silk cocoons. This extract is composed of both carotenoids and flavonoids. Many of these compounds are composed of substances of poor aqueous solubility. Thus, this study focused on the development of a carrier system created from biocompatible and biodegradable materials to improve the biological activity of NS extracts. Accordingly, NS was incorporated into human serum albumin template particles with MnCO3 (NS-HSA MPs) by loading NS into the preformed HAS-MnCO3 microparticles using the coprecipitation crosslinking dissolution technique (CCD-technique). After crosslinking and template dissolution steps, the NS loaded HSA particles are negatively charged, have a size ranging from 0.8 to 0.9 µm, and are peanut shaped. The degree of encapsulation efficiency ranged from 7% to 57% depending on the initial NS concentration and the steps of adsorption. In addition, NS-HSA MPs were taken up by human lung adenocarcinoma (A549 cell) for 24 h. The promotion of cellular uptake was evaluated by flow cytometry and the results produced 99% fluorescent stained cells. Moreover, the results from CLSM and 3D fluorescence imaging confirmed particle localization in the cells. Interestingly, NS-HSA MPs could not induce inflammation through nitric oxide production from macrophage RAW264.7 cells. This is the first study involving the loading of non-sericin extracts into HSA MPs by CCD technique to enhance the bioavailability and biological effects of NS. Therefore, HSA MPs could be utilized as a carrier system for hydrophobic substances targeting cells with albumin receptors.

Hemoglobin-based oxygen carriers (HBOCs) represent a propitious type of blood substitute to transport oxygen throughout the body while acting as a carrier in biomedical applications. However, HBOCs in blood are recognized and rapidly scavenged by the body’s innate immune systems. To overcome this problem, HBOCs require a surface modification that provides protection against detection and elimination in order to prolong their circulation time after administration. In this study, we investigated different surface modifications of hemoglobin submicron particles (HbMPs) by double/triple precipitation, as well as by adsorption of human serum albumin (HSA), hyaluronic acid (HA), and pluronic (Plu) to discover how diverse surface modifications influence the oxygen binding capacity and the binding of anti-hemoglobin (Hb) antibodies, immunoglobulin G (IgG), and haptoglobin (HP) to HbMPs. The particle size and zeta potential of the six types of HbMP modifications were analyzed by zeta sizer, confocal laser scanning microscopy, and transmission electron microscopy (TEM), and were compared to the unmodified HbMPs. The results revealed that all surface-modified HbMPs had a submicron size with a negative charge. A slight decrease in the oxygen binding capacity was noticed. The specific binding of anti-Hb antibodies, IgG, and HP to all surface-modified HbMPs was reduced. This indicates a coating design able to protect the particles from detection and elimination processes by the immune system, and should lead to a delayed clearance and the required and essential increase in half-life in circulation of these particles in order to fulfill their purpose. Our surface modification method reflects a promising strategy for submicron particle design, and can lead the way toward novel biomedical applications.