Synergies between micropreparative high-performance liquid chromatography and an instrumental optical biosensor

The recent development of an automated surface plasmon resonance technology for the measurement of biomolecular interactions (Pharmacia BIAcore) has provided new opportunities for the detection and analysis of protein-protein interactions. In the BIAcore, detection is based on changes in surface plasmon resonance which are monitored optically. Changes in surface plasmon resonance correspond to changes in surface concentration of macromolecules and can be monitored in real time.

We have found that the detection sensitivity obtainable with this technology (ng/ml concentrations of specific ligands are readily detectable for many applications) is complementary “in a bidirectional manner” to micropreparative HPLC. Thus micropreparative HPLC may be used to purify and characterise reagents for the biosensor, whilst the biosensor may be used to define chromatographic parameters such as elution conditions for affinity chromatography or serve as an affinity detector for fractions obtained during chromatographic purification.

Examples of such applications, including the potential of the biosensor to search for and monitor the purification of unknown ligands for which the target molecule has been identified, are shown. In particular, the use of the biosensor to monitor the purification of soluble epidermal growth factor receptor from A431 cell conditioned media is demonstrated.     

Commutator-finite orthomodular lattices

The class of orthomodular lattices which have only finitely many commutators is investigated. The following theorems are proved: contains the block-finite orthomodular lattices. Every irreducible element of is simple. Every element of is a direct product of a Boolean algebra and finitely many simple orthomodular lattices. The irreducible elements of which are modular, or are M-symmetric with at least one atom, have height two or less.     

Sub-microgram-scale analysis by coulometry at controlled potential

The considerations which determine the success of controlled-potential coulometric analyses on the submicrogram level are discussed, and it is shown that the ultimate sensitivity of the method is governed by the accuracy with which the requisite background corrections, especially that for the charging quantity of electricity, can be determined. A method for the coulometric determination of zinc based on these considerations is shown to contain a limiting uncertainty of ± 0.2 mμF_y, so that as little as 0.07 μg of zinc can be determined within about ± 10%, while quantities of zinc exceeding about 10 μg can be determined with an accuracy and precision of ± 0.1% or better. Possible techniques for the further extension of controlled-potential coulometric analysis into the millimicrogram range are briefly discussed.     

Multi-iron wells-Dawson heteropolytungstates. Electrochemical probing of siderophoric behavior in sandwich-type complexes

The demetalation process of 10 multi-iron Wells-Dawson polyoxometalates is studied by cyclic voltammetry and controlled potential coulometry. Eight sandwich-type complexes (alphaalphaalphaalpha-Na(16)[(NaOH(2))(2)(Fe(III))(2)(X(2)W(15)O(56))(2)], alphaalphabetaalpha-Na(14)[(NaOH(2))(Fe(III)OH(2))(Fe(III))(2)(X(2)W(15)O(56))(2)], alphabetabetaalpha-Na(12)[(Fe(III)OH(2))(2)(Fe(III))(2)(X(2)W(15)O(56))(2)], and alphabetabetaalpha-Na(14)[(Mn(II)OH(2))(2)(Fe(III))(2)(X(2)W(15)O(56))(2)] (where X = P(V) or As(V))) and two monomeric complexes (alpha-Na(11)[(P(2)(Fe(III)Cl)(2)(Fe(III)OH(2))W(15)O(59))] and alpha-Na(11)[(As(2)(Fe(III)Cl)(2)Fe(III)OH(2))W(15)O(59))]) were selected for this study. All 10 complexes show Fe(III) waves which are well-separated from the redox activity of the W(VI) centers. At room temperature and under mild conditions, iron release from the complexes is observed upon reduction of the Fe(III) centers. This release is controlled by the ionic strength of the medium, the nature and concentration of the anions present in the supporting electrolyte, and by the pH of the solution. This behavior parallels those described for most siderophores which depend on the same parameters.     

The s-p bonded representatives of the prominent BaAl4 structure type: a case study on structural stability of polar intermetallic network structures

This work presents a detailed, combined experimental and theoretical study on the structural stability of s-p bonded compounds with the BaAl4 structure type (space group I4/mmm, Z = 2) as part of a broad program to investigate the complex questions of structure formation and atomic arrangements in polar intermetallics. From ab initio calculations employing pseudopotentials and a plane wave basis set, we extracted optimized structural parameters, binding energies, and the electronic structure of the systems AeX(III)4, AeX(II)2X(IV)2, AeX(II)2X(III)2 (Ae = Ca, Sr, Ba; X(II) = Mg, Zn; X(III) = Al, Ga; X(IV) = Si, Ge). For all systems we found a pronounced pseudo-gap in the density of states separating network X42- bonding from antibonding electronic states that coincides with the Fermi level for an electron count of 14 electrons per formula unit, the optimum value for stable BaAl4-type polar intermetallics. However, the synthesis and structural characterization (from X-ray single crystal and powder diffraction data) of the new compounds AeZn2-Al2+, AeZn2-deltaGa2+delta (Ae = Ca, Sr, Ba; delta = 0-0.2) and AeMg0.9Al3.1, AeMg1.7Ga2.3 (Ae = Sr, Ba) manifested that electron deficiency is rather frequent for BaAl4-type polar intermetallics. The site preference for different "X" elements in the ternary systems was quantified by calculating "coloring energies", which, for some systems, was strongly dependent on the size of the electropositive Ae component. The Ae2+ cations decisively influence the nearest neighbor distances in the encapsulating polyanionic networks X4(2-) and the structures of these networks are surprisingly flexible to the size of the Ae component without changing the overall bonding picture. A monoclinically distorted variant of the BaAl4 structure occurs when the cations become too small for matching the size of encapsulating X4(2-) cages. An even larger size mismatch leads to the formation of the EuIn4 structure type.     

Spectrophotometric determination of pentaerythritol tetranitrate (PETN) in waste water from lead styphnate primer plants

An accurate spectrophotometric method is proposed for the determination of pentaerythritol tetranitrate (PETN) in waste water from lead styphnate primer plants by use of phenoldisulphonic acid. The waste water is filtered through a sintered glass crucible and the PETN is determined in the filtrate and the residue. In the determination of PETN in the filtrate, sodium hydroxide is added and the PETN is extracted with methylene chloride (in alkaline solution, styphnate and TNT are not extracted). The methylene chloride solution is then evaporated to dryness, the residue is treated with a solution of phenoldisulphonic acid in sulphuric acid, water and ammonia are added, and the yellow colour is measured. In the determination of PETN in the residue, the PETN is dissolved in acetone, an aliquot of the acetone solution is treated with water and sodium hydroxide, the PETN is extracted with methylene chloride and the colour is developed as above. Various factors affecting the determination were investigated. The solubility of PETN in water was studied.     

A photoacoustic study of the collisional deactivation of CO2 by N2, CO and O2 between 160 and 375 K

A photoacoustic method is used under such experimental conditions that the (01¹0) level of CO₂ gas is not in equilibrium with the other vibrational levels. The rate constants κ′₁₀ characterizing the CO₂ (01¹0) collisional deactivation by N₂, CO and O₂ are measured directly.     

Flow-through particles for the high-performance liquid chromatographic separation of biomolecules: perfusion chromatography

This paper reports a new technique for reducing resistance to stagnant mobile phase mass transfer without sacrificing high adsorbent capacity or necessitating extremely high pressure operation. The technique involves the flow of liquid through a porous chromatographic particle, and has thus been termed "perfusion chromatography". This is accomplished with 6000-8000 A pores which transect the particle. Data from electron microscopy, column efficiency, frontal analysis and theoretical modelling all suggest that mobile phase will flow through these large pores. In this manner, solutes enter the interior of the particles through a combination of convective and diffusional transport, with convection dominating for Peclet numbers greater than one. The implications of flow through particles on bandspreading, resolution and dynamic loading capacity are examined. It is shown that the rate of solute transport is strongly coupled to mobile phase velocity such that bandspreading, resolution of proteins and dynamic loading capacity are unaffected by increases in mobile phase velocity up to several thousand centimeters per hour. The surface area of this very large-pore diameter material is enhanced by using a network of smaller, 500-1500 A interconnecting pores between the throughpores. Scanning electron micrographs show that the pore network is continuous and that no point in the matrix is more than 5000-10,000 A from a through-pore. As a consequence, diffusional path lengths are minimized and the large porous particles take on the transport characteristics of much smaller particles but with a fraction of the pressure drop. Capacity and resolution studies show that these materials bind and separate an amount of protein equivalent to that of conventional high-performance liquid chromatography as well as low performance agarose-based media at greater than 10-100 times higher mobile phase velocity with no loss in resolution.     

A new test of the weak equivalence principle

The Eötvös, Dicke, and Braginski experiments do not rule out the recent suggestion that the weak equivalence principle (WEP) might be violated at intermediate ranges (10^–1 m r 10⁴ m). I briefly discuss the problems inherent in Eötvös-type apparatus in searches for WEP-violating forces (hyperforces) between laboratory masses and suggest an alternative detector free of such problems. The proposed detector is driven by a hyperforce torque at the rotational frequency. If the detector is tuned to this frequency, the signal, enhanced by resonance, may be detected synchronously. I derive the response equations for the detector and discuss how spurious responses due to gravity torques may be suppressed.This essay received the third award from the Gravity Research Foundation for the year 1986-Ed.