Ivo Šafařík

Magnetic Nanoparticles and Biosciences

Summary. Magnetic nanoparticles represent an interesting material both present in various living organisms and usable for a variety of bioapplications. This review paper will summarize the information about biogenic magnetic nanoparticles, the ways to synthesize biocompatible magnetic nano- particles and complexes containing them, and the applications of magnetic nanoparticles in various areas of biosciences and biotechnologies.

Direct determination of total soil carbohydrate content

A direct procedure for the determination of total soil carbohydrate content and a classic determination after acid hydrolysis, both employing the phenol-sulphuric acid method, are compared. The direct procedure enables simultaneous determination of mono-, oligo- and polysaccharides without prior hydrolysis. This procedure is reproducible and takes only a short period of time. The correlation between the proposed method and the classic one with hydrolysis was high (r2=0.9843; n=11; p=0.05).

Batch isolation of hen egg white lysozyme with magnetic chitin

Biospecific magnetic sorbent for lysozyme isolation (magnetic chitin) has been prepared from magnetic chitosan after acetylation with acetic anhydride. The capacity of magnetic chitin was 2.5 mg of lysozyme per 1 ml of sorbent.

Large-scale separation of magnetic bioaffinity adsorbents

Flat magnetic separator was used to separate magnetic bioaffinity adsorbents from litre volumes of suspensions. Both magnetic cross-linked erythrocytes and magnetic chitosan were efficiently separated; at least 95% adsorbent recovery was achieved at maximum flow rate (1680 ml min−1). Using this system low amounts of trypsin were concentrated from large sample volumes using magnetic erythrocytes as affinity adsorbent.

Magnetic Nanoparticles for Biomedicine

Biocompatible materials exhibiting different types of response to external magnetic field have already found many important applications in various areas of biosciences, biotechnology, medicine, environmental technology etc. In most cases they can be described as composite materials, where the magnetic properties are caused by the presence of iron oxides nano- or microparticles. Such materials can be efficiently separated from difficult-to-handle samples and targeted to the desired place, applied as contrast agents for magnetic resonance imaging or used to generate heat during exposure to alternating magnetic field.

Separation of magnetic affinity biopolymer adsorbents in a Davis tube magnetic separator

A Davis tube (a matrix-free, flow-through magnetic separator used mainly in mineral processing) has been tested for separation of magnetic affinity biopolymer adsorbents from larger volumes of suspensions. Both magnetic chitosan and magnetic cross-linked erythrocytes could be efficiently separated from litre volumes of suspensions. Up to 90% adsorbent recovery was achieved under optimised separation conditions.

A spectrophotometric assay for lipase activity utilizing immobilized triacylglycerols.

New substrates for the determination of lipase activity have been developed. Triacylglycerols were immobilized by adsorption on an appropriate carrier or adsorbent yielding a lipase substrate in a powder form. The adsorbed triacylglycerols were easily hydrolyzed by lipases present in a reaction mixture. The released fatty acids were extracted with benzene and converted to the corresponding Cu (II) salts (copper soaps) which were measured spectrophotometrically.

A modified procedure for the preparation of insoluble chromogenic substrates for the determination of proteolytic activity

New insoluble chromogenic substrates for the determination of proteolytic activity were prepared by cross-linking gelatin with glutaraldehyde in the presence of congo red or nigrosin. The prepared substrates could detect approximately 0.4 microgram of trypsin per ml. The spontaneous leakage of dyes in water solutions was negligible.

An insoluble chromolytic substrate for the determination of proteolytic activity

Abstract A new insoluble chromolytic substrate for the determination of proteolytic activity was prepared by immobilization of dyed casein into the structure of polyacrylamide gel. The prepared substrate could detect approximately 0.1 μg of trypsin per ml. The spontaneous leakage of dyed casein molecules in water solutions was negligible.

Improved properties of bovine erythrocyte acetylcholinesterase, isolated by papain cleavage

A simple and rapid procedure involving papain cleavage of the membrane anchor was used to isolate membrane-bound acetylcholinesterase from bovine erythrocytes. The solubilized enzyme was purified 930-fold by ion exchange chromatography and gel filtration. The properties of the papain-cleaved acetylcholinesterase were compared with those of a commercial acetylcholinesterase, solubilized from the erythrocyte membranes by detergents. Cleavage of the membrane anchor eliminated dimer aggregation, caused a pH shift in thermal stability and resulted in increased stability in organic solvents. Bovine serum albumin, used as stabilizer of the commercial enzyme preparation, increased the thermal stability but concomitantly decreased the activity of acetylcholinesterase at pH 6-8. The improved stability of the cleaved acetylcholinesterase, especially in organic solvents, may enhance the biosensor performance of the enzyme.