Isothermal biocalorimetry. A status report.

A brief status report is given for the field of isothermal calorimetry applied to systems of biological interest. These include, for instance, water and simple model compounds, biopolymers, living cells, tissues and animals. Examples of recent thermodynamic work as well as investigations of a more general analytical nature are discussed. Special attention is given to recent developments in instrumentation and methodology.     

Birkhoff’s Polytope and Unistochastic Matrices,N = 3 and N = 4

The set of bistochastic or doubly stochastic N×N matrices is a convex set called Birkhoff’s polytope, which we describe in some detail. Our problem is to characterize the set of unistochastic matrices as a subset of Birkhoff’s polytope. For N=3 we present fairly complete results. For N=4 partial results are obtained. An interesting difference between the two cases is that there is a ball of unistochastic matrices around the van der Waerden matrix for N=3, while this is not the case for N=4.     

Fourth-order accurate one-step integration methods with large imaginary stability limits

One-step integration methods of fourth-order accuracy using an odd number of function evaluations K, to solve dy/dt = A · y, are proposed. These methods have an imaginary stability limit \documentclass{article}\pagestyle{empty}\begin{document}$ S_{1\;} = \sqrt {(K - 1)^2 - 4} $\end{document}. In the case K = 5 the Kutta-Merson method results.     

Use of hydrogen-sensitive Pd-MOS materials in biochemical analysis

A metal oxide semiconductor sensitive to hydrogen was used to monitor H₂ production from immobilizedClostridium acetobutylicum cells and from immobilized hydrogenase on a continuous basis.     

A versatile polypeptide platform for integrated recognition and reporting: affinity arrays for protein-ligand interaction analysis

A molecular platform for protein detection and quantification is reported in which recognition has been integrated with direct monitoring of target-protein binding. The platform is based on a versatile 42-residue helix-loop-helix polypeptide that dimerizes to form four-helix bundles and allows site-selective modification with recognition and reporter elements on the side chains of individually addressable lysine residues. The well-characterized interaction between the model target-protein carbonic anhydrase and its inhibitor benzenesulfonamide was used for a proof-of-concept demonstration. An affinity array was designed where benzenesulfonamide derivatives with aliphatic or oligoglycine spacers and a fluorescent dansyl reporter group were introduced into the scaffold. The affinities of the array members for human carbonic anhydrase II (HCAII) were determined by titration with the target protein and were found to be highly affected by the properties of the spacers (dissociation constant Kd=0.02-3 microM). The affinity of HCAII for acetazolamide (Kd=4 nM) was determined in a competition experiment with one of the benzenesulfonamide array members to address the possibility of screening substance libraries for new target-protein binders. Also, successful affinity discrimination between different carbonic anhydrase isozymes highlighted the possibility of performing future isoform-expression profiling. Our platform is predicted to become a flexible tool for a variety of biosensor and protein-microarray applications within biochemistry, diagnostics and pharmaceutical chemistry.     

A comparison of some flame atomic absorption spectrometric methods for the determination of cadmium in fertilizers and related materials

A liquid-liquid extraction procedure for cadmium in phosphate rock and phosphate-based fertilizers is described. Extraction with tri-n-octylamine and potassium iodide into methyl isobutyl ketone is shown to be simpler and less prone to interferences than the more commonly used methods based on ammonia pyrrolidine-dithiocarbamate. Iron interest strongly in the dithiocarbamate method. Addition of ascorbic acid, and extraction at low pH allows the dithiocarbamate method to operate up to 0.15 g Fe l^?1, while the amine method tolerates at least 0.2 g Fe l^?1. Direct determination is suitable on samples with a sufficiently high cadmium content. The amine extraction method has been applied successfully at an apatite containing only 0.03 mg kg^?1 cadmium.     

Investigation of sera from various species by using lectin affinity arrays and scanning ellipsometry

Serum proteins of different species and of different human blood groups exhibit various protein glycosylation patterns. Sera from human, pig, sheep and guinea pig have been applied to a panel of eight different lectins immobilized on a gold wafer. The biorecognition has been evaluated with scanning ellipsometry and the two-dimensional matrices obtained have been treated with image analysis and MVDA for evaluation. The results showed a clear difference in protein binding pattern between the different species and thereby separation of the different sera could be made. Dendograms indicate that human and pig sera are the most related of the four different sera investigated.     

Biosensor discrimination of meat juice from various animals using a lectin panel and ellipsometry

In this work, simple microcontact printed gold-wafers were used to make a lectin panel for investigation and discrimination of different meat juices from fresh meat of cattle, chicken, pig, cod, turkey and lamb. Seven different lectins were thus attached to gold surfaces using the streptavidin-biotin method. Lectins recognize and bind specifically to carbohydrate structures present on different proteins. The biorecognition was evaluated with null ellipsometry and the data obtained was related to an internal standard of lactoferrin. The data was evaluated with multivariate data analysis techniques to identify possible discrimination or grouping of data. Scanning ellipsometry was used for visualization of the binding pattern of the lectins and the meat juice proteins. The two-dimensional images obtained could be used to visualize the protein distribution, furthermore, to exclude anomalies. The results showed that the different meat juices from the six different species: cattle, chicken, pig, cod, turkey and lamb could be discriminated from each other. The results showed to be more repetitive for the mammalian meat juices. Using a simple model based on an artificial neuronal net, it was also possible to classify meat juices from the mammals investigated.     

Microcalorimetric techniques for the investigation of living plant materials

Microcalorimetric investigations of living cellular systems have been conducted on small animals, cells and tissues from animals and plants and on microbial systems, but it is only recently that more systematic work has begun on plant materials. Isothermal experiments together with temperature scanning experiments have been performed. In most cases simple static ampoules have been used as calorimetric vessels and practically all calorimetric investigations reported on plant systems have been conducted under dark conditions. It is felt that some microcalorimetric techniques currently used in studies of microbial and animal systems, and in other areas, could be usefully applied more generally in the plant field: stirred injection (titration) vessels, gas and liquid perfusion vessels and different photocalorimetric methods.