Magnetophoretic and spectral characterization of oxyhemoglobin and deoxyhemoglobin: Chemical versus enzymatic processes.

A new method for hemoglobin (Hb) deoxygenation, in suspension or within red blood cells (RBCs) is described using the commercial enzyme product, EC-Oxyrase®. The enzymatic deoxygenation method has several advantages over established deoxygenation methodologies, such as avoiding side reactions that produce methemoglobin (metHb), thus eliminating the need for an inert deoxygenation gas and airtight vessel, and facilitates easy re-oxygenation of Hb/RBCs by washing with a buffer that contains dissolved oxygen (DO). The UV-visible spectra of deoxyHb and metHb purified from human RBCs using three different preparation methods (sodium dithionite [to produce deoxyHb], sodium nitrite [to produce metHb], and EC-Oxyrase® [to produce deoxyHb]) show the high purity of deoxyHb prepared using EC-Oxyrase® (with little to no metHb or hemichrome production from side reactions). The oxyHb deoxygenation time course of EC-Oxyrase® follows first order reaction kinetics. The paramagnetic characteristics of intracellular Hb in RBCs were compared using Cell Tracking Velocimetry (CTV) for healthy and sickle cell disease (SCD) donors and oxygen equilibrium curves show that the function of healthy RBCs is unchanged after EC-Oxyrase® treatment. The results confirm that this enzymatic approach to deoxygenation produces pure deoxyHb, can be re-oxygenated easily, prepared aerobically and has similar paramagnetic mobility to existing methods of producing deoxyHb and metHb.

Novel annulene dications from methylated [2.2]metacyclophane monoenes and [e]-ring benzannelated dimethyldihydropyrene.

Tetramethyl- and hexamethyl-substituted [2.2]metacyclophane monoenes (10 and 11) are transformed into their corresponding trans-dimethyldihydroethanophenanthrenium dications (14(2+) and 15(2+)) in FSO(3)H x SbF(5) (4:1) and FSO(3)H x SbF(5) (1:1) with SO(2)ClF or SO(2) as the solvent; these 10 pi-dications are equivalent to the C-4/C-5 diprotonated dications of the 2,7-dimethyl derivative of trans-DMDHP, 3a. The trans-12c,12d-dimethyl-12c,12d-dihydrobenzo[e]pyrene (6) reacts with FSO(3)H/SO(2)ClF under surprisingly mild conditions to give initially a persistent diprotonated dication (6H(2)(2+)) and, subsequently, the oxidation dication (6(2+)); the 6(2+):6H(2)(2+) ratio reaches 4:1 after 1 week at low temperature. Protonation of the anti-metacyclophane (13) was also examined. Charge delocalization mode and tropicity in the resulting dications are gauged via detailed NMR studies at 500 MHz.

Dehydrobenzoannulene-dimethyl-dihydropyrene hybrids: model systems for the synthesis of molecular aromatic probes.

Synthesis of two novel dehydrobenzoannulene-dimethyldihydropyrene (DBA-DDP) hybrids has been achieved using a Pd/Cu cross-coupling strategy.

Backbone photochromic polymers containing the dimethyldihydropyrene moiety: toward optoelectronic switches.

The design and synthesis of the first electrically conducting main chain photochromic conjugated polymer incorporating dimethyldihydropyrene is reported, as is the demonstration of the corresponding repeat unit as an optoelectronic redox switch.

Fast and reliable QRS alignment technique for high-frequency analysis of signal-averaged ECG.

The process of QRS alignment as required in signal-averaged ECG can impose serious limitations on the spectral range of the signal output. This effect depends basically on the particular alignment technique being used and on the level and type of noise present in the recorded ECG. In clinical studies where a wide-band (1000 Hz) ECG averager is required, the conventional QRS alignment technique, based on maximum coherence matching (MCM) with a template beat, may not perform consistently well. An alternative QRS alignment technique based on the accurate detection of a single fiducial point (SFP) in the bandpass filtered (3-30 Hz) QRS complex was developed. Using computer simulation methods, a comparative assessment of the frequency bandwidths (3 dB points) offered by both MCM and SFP techniques as a function of noise level (15-100 muRMS) and type (EMG and 50 Hz interference), was carried out. The results of the comparative assessment indicated a better performance by the SFP technique in all cases of noise. Hence, the SFP technique would perform more reliably for high-frequency analysis of a noisy ECG, especially when 50 Hz interference is high. Furthermore, SFP is considerably faster than MCM (about four times) when implemented digitally, and its analogue realisation is feasible. The SFP technique is suitable for late-potential analysis in the signal-averaged ECG.

Cellular automation model of ventricular conduction.

A computer simulation of ventricular conduction is presented with a discrete element ventricular conduction model. The model is shown to be capable of demonstrating normal ventricular rhythm, including QRS and T-waves, plus a variety of rhythm disturbances including missed beats, electrical alternans and self-sustaining instability with arrhythmias. The morphology of the range of alternans produced bears a strong resemblance to certain clinically recognised arrhythmias.

Power spectral analysis of the electrocardiogram in diabetic children.

In recent years it has been shown that alteration in heart rate variability can be used for the objective assessment of autonomic function in adult diabetic patients. Using a microcomputer based system for on-line monitoring and analysis of heart rate variability by power spectral analysis, 100 children with Type 1 (insulin-dependent) diabetes mellitus were studied. A highly significant negative correlation was identified between heart rate variability and duration of diabetes (r = -0.88, p less than 0.0001). The mean heart rate variability in patients with diabetes of duration 3 years or more was significantly lower in comparison to age-matched control subjects. A highly significant negative correlation was evident between heart rate variability and mean HbA1 in patients with duration of diabetes of 5 years or more. A mean HbA1 over 10% during this period was associated with the greatest reduction in heart rate variability. Serial measurements of heart rate variability in diabetic children may be of predictive value prior to the onset of symptomatic neuropathy.

Cellular automaton model of ventricular fibrillation.

A theoretical analysis of ventricular fibrillation and the requirements for fibrillation are performed using a discrete element neighborhood (cellular automaton) model of ventricular conduction. The model is configured as a 2500 element rectangular grid on the surface of a cylinder. It is shown that vulnerability to fibrillation is strongly influenced by excited state duration which primarily determines the nature of the underlying reentry activity. As excited state duration is increased fibrillation changes from "coarse" macroreentrant activity to the more chaotic "fine" fibrillation sustained by multiple wavelets of microreentry. In general, defibrillation is achieved by a stimulus strong enough to depolarize the majority of relative refractory elements. The threshold for defibrillation is increased for the more irregular microreentrant fibrillation.

Electrical stimulation of cultured myocardial cells.

This paper reports on the use of a cardiac tissue monolayer model in the investigation of some of the fundamental electrical properties of cardiac muscle. The response of cardiac tissue to increasing levels of electrical stimulus is investigated and the strength-duration curve for rectangular waveform stimulation is measured. An experimental protocol is established which provides for uniform field stimulation of the cells, and allows the cellular response (contraction) to be recorded.

A gas-flow cryostat for use in freeze-quench studies: design and application to discontinuous pre-steady-state spectral analyses.

A design for an improved freeze-quench apparatus is presented. A freeze-quench apparatus has two main parts: the apparatus which mixes the reactants and after a set time sprays the reacting mixture into the quencher, and the quenching apparatus itself. The quenching apparatus is the novel feature of our design and it comprises a gas-flow cryostat mounted directly onto a liquid nitrogen storage Dewar. The gas flow maintains the temperature of a small volume of isopentane, which acts as the freeze-quench agent, at -140 degrees C. (The apparatus can maintain the quenching temperature over the range -190 degrees C to room temperature.) Because of the small size of the cryostat and the much reduced volume of solvent used by this method it is more convenient to use than its predecessors, can be used in the open laboratory, and is safer. Our apparatus is designed for application to electron paramagnetic resonance studies but could be easily modified for use with other spectroscopic techniques.

A computer model of re-entry in cardiac tissue.

This paper describes the use of a computer model to investigate the nature of impulse propagation and the mechanisms of electrical instability which precipitate ventricular arrhythmias. The model provides the opportunity for a systematic analysis of the conditions of cardiac conduction under controlled conditions. From the simulation studies performed it is found that the most critical parameter which influences re-entry circuit formation is the ratio of exited to refractory duration.

Real time analysis of Valsalva's maneuver.

This paper describes a real-time system for the evaluation of Valsalva's Maneuver. The computer software is menu-driven and produces the parameters of linear and nonlinear models which can be used to describe the different phases of the Valsalva Maneuver. The system is routinely in use as a means of assessing baroreflex function during the Valsalva Maneuver following drug administration.

Portable system for the measurement of heart rate.

We describe a simple RR interval recording system; it does not involve an analogue recording of ECG data on magnetic tape and does not therefore suffer from the inherent problem of tape speed variability on playback, which is a frequent cause of inaccuracies in RR interval timing. The system operates in both recording and playback modes and records of up to 60 minutes duration can be obtained without operator intervention. Its accuracy with respect to RR interval is not less than 99.5%.

Induction of DNA-DNA cross-link formation in human cells by various psoralen derivatives.

Furocoumarin-induced DNA damage, monoadducts, and cross-links were measured in normal human, xeroderma pigmentosum, and Fanconi's anemia cells after exposure to near-UV (356 nm). At similar concentrations and near-UV doses, photoaddition by 8-methoxypsoralen was twice that by angelicin and the substitution of bromodeoxyuridine for thymidine in one strand of DNA did not alter the binding. The rate of cross-linking by 8-methoxypsoralen was twice that of 5-methoxypsoralen. Low frequencies of cross-links were detected from angelicin and 3-carbethoxypsoralen but none were detected from 5-geranoxypsoralen at concentrations up to 25 micrograms/ml and near-UV doses up to 45,000 J/m2.

The development of monitoring techniques in mobile coronary care.

Mobile coronary care was first introduced by Pantridge and Geddes at the Royal Victoria Hospital in 1966. Traditional emergency equipment i.e. defibrillation and monitors were unsuitable for application in this environment. In the succeeding years the defibrillator was made highly portable, and significantly reduced in size. Specialized monitoring apparatus was also developed to provide display and recording facilities to meet the special needs of the mobile coronary care unit. However, current display systems utilize the cathode ray oscilloscope, which does not lend itself to miniaturization, is fragile, and cannot be placed at close proximity to the patient. A flat screen electroluminescent matrix of 256 X 128 dots has been developed as a physiological data display. The panel achieves a line density of approximately 83 per inch and with a time compressed memory and interpolated drive technique provides an acceptable display.

Properties of colony-stimulating and inhibiting activities of chicken serum.

Serum obtained from young chicks stimulated with endotoxin contained elevated levels of colony-stimulating activity (CSA). This CSA was partially characterized. Incubation at 65 degrees C (60 min) had no effect on the ability of the CSA to support colony formation by nonadherent cells but reduced 3-4 fold its ability to support colony formation by unfractionated marrow cells. These data suggest that most of the serum activity is due to a heat-labile, adherent cell-dependent factor which does not directly affect the nonadherent granulocyte/monocyte colony-forming cell (GM-CFC). The ability of serum to support colony formation was increased by chloroform extraction. This was evidently due to the presence of inhibitory lipoproteins. The lipoprotein fraction of pigeon serum was found to directly block colony formation. The CSA for nonadherent cells migrated as one peak in the postalbumin region during zone electrophoresis. When electrofocused, heterogeneity was detected but all activity was associated with molecules with isoelectric points between pH 4 and 5. Following sequential fractionation by chloroform extraction, ultrafiltration, zone electrophoresis, isoelectric focusing and gel filtration (Ultrogel AcA 44), the activity was separated from most of the remaining protein. One peak of activity was obtained at the excluded volume.