Investigation of a circannual rhythm in nasopharyngeal airway patency in children

We have studied nasopharyngeal adenoids on standardized cephalometric radiograms of 25 children in each season. While individual subjects showed changes ranging from 1-17% throughout the year, a synchronized circannual rhythm could be described for the girls only. Similar changes were observed in boys, although a synchronized group rhythm could not be ascertained. No clear explanation for this apparent sexual dimorphism is known at this time, but different hormonal influences are likely. Evaluation of the nasopharyngeal airway (NPA) patency is thus affected by the season in which a cephalogram is taken. It should be noted that due to the described circannual rhythmicity, a single determination of the adenoid masses may be an insufficient and misleading indicator of the NPA obstruction. Etiology and clinical significance of these rhythmic changes need to be explored further.     

The involvement of iron and ubiquinone in electron transfer reactions mediated by reaction centers from photosynthetic bacteria.

Reaction centers from Rhodopseudomonas sphaeroides strain R-26 were prepared with varying Fe and ubiquinone (Q) contents. The photooxidation of P-870 to P-870+ was found to occur with the same quantum yield in Fe-depleted reaction centers as in control samples. The kinetics of electron transfer from the initial electron acceptor (I) to Q also were unchanged upon Fe removal. We conclude that Fe has no measurable role in the primary photochemical reaction. The extent of secondary reaction from the first quinone acceptor (QA) to the second quinone acceptor (QB) was monitored by the decay kinetics of P-870+ after excitation of reaction centers with single flashes in the absence of electron donors, and by the amount of P-870 photooxidation that occurred on the second flash in the presence of electron donors. In reaction centers with nearly one iron and between 1 and 2 ubiquinones per reaction center, the amount of secondary electron transfer is proportional to the ubiquinone content above one per reaction center. In reaction centers treated with LiClO4 and o-phenanthroline to remove Fe, the amount of secondary reaction is decreased and is proportional to Fe content. Fe seems to be required for the secondary reaction. In reaction centers depleted of Fe by treatment with SDS and EDTA, the correlation between Fe content and secondary activity is not as good as that found using LiClO4. This is probably due in part to a loss of primary photochemical activity in samples treated with SDS; but the correlation is still not perfect after correction for this effect. The nature of the back reaction between P-870+ and Q-B was investigated using stopped flow techniques. Reaction centers in the P-870+ Q-B state decay with a 1-s half-time in both the presence and absence of o-phenanthroline, an inhibitor of electron transfer between Q-B and QB. This indicates that the back reaction between P-870+ and Q-A is direct, rather than proceeding via thermal repopulation of Q-A. The P-870+ Q-B state is calculated to lie at least 100 mV in free energy below the P-870+ Q-A state.     

Nanosecond fluorescence from chromatophores of Rhodopseudomonas sphaeroides and Rhodospirillum rubrum

Single-photon counting techniques were used to measure the fluorescence decay from Rhodopseudomonas sphaeroides and Rhodospirillum rubrum chromatophores after excitation with a 25-ps, 600-nm laser pulse. Electron transfer was blocked beyond the initial radical-pair state (PF) by chemical reduction of the quinone that serves as the next electron acceptor. Under these conditions, the fluorescence decays with multiphasic kinetics and at least three exponential decay components are required to describe the delayed fluorescence. Weak magnetic fields cause a small increase in the decay time of the longest component. The components of the delayed fluorescence are similar to those found previously with isolated reaction centers. We interpret the multi-exponential decay in terms of two small (0.01-0.02 eV) relaxations in the free energy of PF, as suggested previously for reaction centers. From the initial amplitudes of the delayed fluorescence, it is possible to calculate the standard free-energy difference between the earliest resolved form of PF and the excited singlet state of the antenna complexes in R. rubrum strains S1 and G9. The free-energy gap is found to be about 0.10 eV. It also is possible to calculate the standard free-energy difference between PF and the excited singlet state of the reaction center bacteriochlorophyll dimer (P). Values of 0.17 to 0.19 eV were found in both R. rubrum strains and also in Rps. sphaeroides strain 2.4.1. This free-energy gap agrees well with the standard free-energy difference between PF and P determined previously for reaction centers isolated from Rps. sphaeroides strain R26. The temperature dependence of the delayed fluorescence amplitudes between 180 K and 295 K is qualitatively different in isolated reaction centers and chromatophores. However, the temperature dependence of the calculated standard free-energy difference between P* and PF is similar in reaction centers and chromatophores of Rps. sphaeroides. The different temperature dependence of the fluorescence amplitudes in reaction centers and chromatophores arises because the free-energy difference between P* and the excited antenna is dominated by the entropy change associated with delocalization of the excitation in the antenna. We conclude that the state PF is similar in isolated reaction centers and in the intact photosynthetic membrane. Chromatophores from Rps. sphaeroides strain R-26 exhibit an anomalous fluorescence component that could reflect heterogeneity in their antenna.     

System for flow sorting chromosomes on the basis of pulse shape

Sorting on the basis of the complex features resolved by chromosome slit-scan analysis requires rapid and flexible pulse shape acquisition and processing for determining sort decisions before droplet breakoff. Fluorescence scans of chromosome morphology contain centromeric index and banding information suitable for chromosome classification, but these scans are often characterized by variability in length and height and require sophisticated data processing procedures for identification. Setting sort criteria on such complex morphological data requires digitization and subsequent computation by an algorithm tolerant of variations in overall pulse shape. We demonstrate here the capability to sort individual chromosomes based on their morphological features measured by slit-scan flow cytometry. To do this we have constructed a sort controller capable of acquiring an 128 byte chromosome waveform and executing a series of numerical computations resulting in an area-based centromeric index sort decision in less than 2 ms. The system is configured in a NOVIX microprocessor, programmed in FORTH, and interfaced to a slit-scan flow cytometer data acquisition system. An advantage of this configuration is direct control over the machine state during program execution for minimal processing time. Examples of flow sorted chromosomes are shown with their corresponding fluorescence pulse shapes.     

Excited states of photosynthetic reaction centers at low redox potentials

In preparations of photochemical reaction centers from Rhodopseudomonas spheroides R-26, lowering the redox potential so as to reduce the primary electron acceptor prevents the photochemical transfer of an electron from bacteriochlorophyll to the acceptor. Measuring absorbance changes under these conditions, we found that a 20-ns actinic flash converts the reaction center to a new state, P^F, which then decays with a half-time that is between 1 and 10 ns at 295 °K. At 25 °K, the decay half-time is approx. 20 ns. The quantum yield of state P^F appears to be near 1.0, both at 295 and at 15 °K. State P^F could be an intermediate in the photochemical electron-transfer reaction which occurs when the acceptor is in the oxidized form.Following the decay of state P^F, we detected another state, P^R, with a decay half-time of 6 μs at 295 °K and 120 μs at 15 °K. The quantum yield of state P^R is approx. 0.1 at 295 °K, but rises to a value nearer 1.0 at 15 °K. The kinetics and quantum yields are consistent with the view that state P^R forms from P^F. State P^R seems likely to be a side-product, rather than an intermediate in the electron-transfer process.The decay kinetics indicate that state P^F cannot be identical with the lowest excited singlet state of the reaction center. One of the two states, P^F or P^R, probably is the lowest excited triplet state of the reaction center, but it remains unclear which one.     

Fast changes of enthalpy and volume on flash excitation of Chromatium chromatophores

We have used a capacitor microphone transducer to measure volume changes in suspensions of Chromatium chromatophores 100 μs to 20 ms after flash excitation. Volume changes in photochemical reactions can arise both from volume differences between reactants and products, and from enthalpy changes which heat or cool the solution. By measuring the volume changes at two temperatures, one can resolve the total changes into their two components.     

Exciton interactions in dimers of bacteriochlorophyll and related molecules

Formalisms are developed for calculating the absorption wavelengths, dipole strengths and rotational strengths for dimers of bacteriochlorophyll and related molecules. The expressions explicitly consider the mixing of bacteriochlorophyll's four main excited states (Q_y, Q_x, B_x and B_y) in the ground and excited states of the dimer. This mixing must be considered in order to account for the hyperchromism and nonconservative circular dichroism found experimentally in oligomers of bacteriochlorophyll and bacteriopheophytin. The spectroscopic properties of the eight absorption bands of a bacteriopheophytin dimer are calculated as functions of the geometry of the dimer. The importance of the mixing of nondegenerate excited states, and of the mixing of doubly-excited states into the dimer's ground state, is evaluated by comparisons with calculations in which these phenomena are neglected. Structures for bacteriopheophytin dimers are found for which most of the calculated spectroscopic properties are consistent with the properties seen experimentally. Possible explanations are considered for the remaining discrepancies between the calculated and observed properties.     

Singlet-triplet fusion in Rhodopseudomonas sphaeroides chromatophores. A probe of the organization of the photosynthetic apparatus

Chromatophores from various strains of Rhodopseudomonas sphaeroides were excited with laser flashes lasting about 20 ns. Fluorescence from the antenna bacteriochlorophyll of the photosynthetic apparatus was measured both during the laser flash, and during a weak Xe flash following the laser flash. Strong laser flashes caused severe quenching of the fluorescence, which could be correlated with the formation of triplet states of the antenna pigments. Triplet states of both BChl and carotenoids acted as quenchers, but bacteriochlorophyll triplets were the more effective of the two. In the double-flash experiments, the reciprocal of the fluorescence yield was proportional to the concentration of triplet quenchers remaining at the time of the second flash. This relationship indicates that singlet excitations can migrate over large domains in the antenna, rather than being restricted by boundaries separating individual reaction centers. Comparisons of chromatophores from different strains and from cells grown under different conditions showed that excitations are concentrated rapidly in the antenna complexes with the longest wavelength absorption bands (B870), and that the migration of excitations to trapping sites is relatively insensitive to the amount of antenna bacteriochlorophyll absorbing at shorter wavelengths (B800–B850). This suggests that the B870 complexes are organized in the membrane so as to interconnect many reaction centers, and that the B800–B850 complexes are arranged peripherally.