A study of artifacts and their removal during forced oscillation of the respiratory system.

Respiratory impedance measured by the forced oscillation technique (FOT) can be contaminated by artifacts such as coughing, vocalization, swallowing or leaks at the mouthpiece. We present a novel technique to detect these artifacts using multilevel discrete wavelet transforms. FOT was performed with artifacts introduced during separate 60 s recordings at known times in 10 healthy subjects. Brief glottal closures were generated phonetically and confirmed by nasopharyngoscopic imaging of the glottis. Artifacts were detected using Daubechies wavelets by applying a threshold to squared detail coefficients from the wavelet transforms of both pressure and flow signals. Sensitivity and specificity were compared over a range of thresholds for different level squared detail coefficients. Coughs could be identified using 1st level detail (cd1) coefficients of pressure achieving 96% sensitivity and 100% specificity while swallowing could be identified using cd2 thresholds of pressure with 95% sensitivity and 97% specificity. Male vocalizations could be identified using cd1 coefficients with 88% sensitivity and 100% specificity. For leaks at the mouthpiece, cd3 thresholds of flow could identify these events with 98% sensitivity and 99% specificity. Thus, this method provided an accurate, easy, and automated technique for detecting and removing artifacts from measurements of respiratory impedance using FOT.

Identifying regions of membrane proteins in contact with phospholipid head groups: covalent attachment of a new class of aldehyde lipid labels to cytochrome c oxidase.

A series of amine-specific reagents based on the benzaldehyde reactive group have been synthesized, characterized, and used to study beef heart cytochrome c oxidase reconstituted in phospholipid bilayers. The series contained three classes of reagents: lipid-soluble phosphodiesters having a single hydrocarbon chain, phospholipid analogues, and a water-soluble benzaldehyde. All reagents were either radiolabeled or spin-labeled or both. The Schiff bases formed by these benzaldehydes with amines were found to be reversible until the addition of the reducing agent sodium cyanoborohydride, whereas attachment of lipid-derived aliphatic aldehydes was not readily reversible in the absence of the reducing agent. The benzaldehyde group provides a convenient method of controlling and delaying permanent attachment to integral membrane proteins until after the reconstitution steps. This ensures that the lipid analogues are located properly to identify amine groups at the lipid-protein interface rather than reacting indiscriminately with amines of the hydrophilic domains of the protein. The benzaldehyde lipid labels attach to cytochrome c oxidase with high efficiency. Typically, 20% of the amount of lipid label present was covalently attached to the protein, and the number of moles of label incorporated per mole of protein ranged from 1 to 6, depending on the molar ratios of label, lipid, and protein. The efficiency of labeling by the water-soluble benzaldehyde was much less than that observed for any of the lipid labels because of dilution effects, but equivalent levels of incorporation were achieved by increasing the label concentration. Electron spin resonance spectra of a nitroxide-containing phospholipid analogue covalently attached to reconstituted cytochrome c oxidase exhibited a large motion-restricted component, which is characteristic of spin-labeled lipids in contact with the hydrophobic surfaces of membrane proteins. The line shape and splittings were similar for covalently attached label and label free to diffuse and contact the protein molecules in the bilayer, providing independent evidence that the coupling occurs at the protein-lipid interface. The distribution of the benzaldehyde reagents attached to the polypeptide components of cytochrome c oxidase was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The labeling pattern observed for the lipid analogues was not affected by the presence of the nitroxide moiety on the acyl chains but was dependent on the molar ratio of labeling reagent to protein.(ABSTRACT TRUNCATED AT 400 WORDS)