A spin-label study of sciatic nerves from quaking, jimpy and trembler mice.

The spin labels, 5-nitroxide stearic acid and 16-nitroxide stearic acid were incorporated into whole sciatic nerves dissected from normal, quaking, jimpy and trembler mice. With 5-nitroxide stearic acid, we have studied the thermal variation of the maximal apparent coupling constant (T) between 0 degrees C and 50 degrees C. Within this range of temperatures, we obtained identical values of 2 T for nerves from normal and jimpy mice, whereas 2 T was smaller for nerves from quaking and trembler mice. With 16-nitroxide stearic acid, composite spectra were recorded, particularly in the high-field range. A line characteristic of myelin was clearly observed in the spectra of nerves from normal and jimpy mice; its intensity was somewhat less in nerves from quaking mice and much less in spectra from trembler mice. A shoulder in the principal highfield line of the spectrum is modified only with nerves from jimpy mice. The results agree well with those obtained by electron microscopy, which reveal normal myelination in nerves from jimpy mice, a slight modification of the myelin from those of quaking mice and a practically complete demyelination in peripheral nerves from trembler mice. However, the structure of the nerves of jimpy mice also seems to be modified at an, as yet, undetermined level.     

X-ray diffraction study of myelin structure in immature and mutant mice

X-ray diffraction patterns were obtained from freshly dissected central and peripheral nerves of quaking, myelin synthesis deficiency ($\text{msd}$), and trembler mutants, as well as immature and adult normal mice. The patterns were compared with respect to strength of myelin diffraction, background scatter level, repeat period, and intensity and linewidth of Bragg reflections. The deficiency of myelin in optic nerves was found to be (in decreasing severity): quaking > immature > trembler ? normal adult; and in sciatic nerves: $\text{trembler > immature > quaking}\text{msd ?}\text{normal}$ adult. Repeat periods about 3 Å less than that for normal adult sciatic myelin were detected in corresponding nerves from immature, quaking, and trembler mice. In some trembler sciatic nerves a second phase having a 190–200 Å period and accounting for about 60% of the total ordered myelin was also evident. Comparison of electron density profiles of membrane units calculated from the repeat periods and diffracted intensities for sciatic myelins indicate structural differences at the molecular level. The main findings are: (1) quaking myelin shows a significant elevation of density in the external protein-water layer between membrane bilayers; (2) the membrane bilayer of immature myelin is ≈ 2 Å thinner than that for normal adult; (3) the membrane bilayer of the more compact phase in trembler myelin is ≈ 5 Å thinner than for normal; and (4) the difference in repeat periods for the two phases present in some of the trembler nerves can be accounted for predominantly by distinct membrane bilayer separations at the external boundary.     

Cellular aspects of tolerance. VII. Inheritance of the resistance to tolerance induction

The half-lives of elimination ($\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$) of ¹³¹I-RGG from the body of normal A or Balb/c animals was much longer than the $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of SJL mice. At all ages, the $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of normal hybrids (A × SJL, SJL × A, Balb/c × SJL) was similar to or longer than that of the A or Balb/c parents. Thus, in terms of the $\text{T}\text{1}\text{2}$ of normal animals, the SJL responsiveness to ¹³¹I-RGG appeared to be a recessive trait. Tolerance could be induced in newborn animals and, in terms of $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$, the degree of unresponsiveness at the age of 6 weeks, was the same in A, Balb/c, A × SJL, and Balb/c × SJL animals but was much shorter in SJL mice. Thus, in neonatally induced tolerance, the duration of tolerance was recessive for the SJL type. The average $\text{T}{}_{\mathrm{<mtext>built1</mtext><mtext>2</mtext>}}$ after tolerance induction in 3-week-old hybrids (A × SJL, SJL × A, Balb/c × SJL) was similar to that of the A or Balb/c parent, but by the 8th and 12th week it approached the average $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of the SJL parent. Comparing 8-week-old hybrids, the average $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ was longest in A × SJL hybrids and identical in SJL × A and Balb/c × SJL mice. An examination of $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ distribution in various 8- and 12-week-old crosses and backcrosses revealed a fairly large proportion of individuals with a $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ which was intermediate between SJL and the other parent. There was a tendency for this number to decrease in 12 weeks as compared to 8-week-old mice. In 8-week-old mice, the number of animals with intermediate $\text{T}{}_{\mathrm{<mtext>built1</mtext><mtext>2</mtext>}}$ was smallest when SJL was the maternal animal [(SJL × A); SJL × (A × SJL); SJL × (SJL × A)]. There was no link between $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of tolerant animals and either the immunoglobulin allotype (MuAl/MuA2) or the C5 eniotype (MuB1 positive/MuB1 negative).     

Asymmetric antagonistic effects of an inhalation anesthetic and high pressure on the phase transition temperature of dipalmitoyl phosphatidic acid bilayers

The phase transition temperature ($\text{T}{}_{\mathrm{<mtext>t</mtext>}}$) of dipalmitoyl phosphatidic acid multilamellar liposomes is depressed 10°C by the inhalation anesthetic methoxyflurane at a concentration of 100 mmol/mol lipid. Application of 100 atm of helium pressure to pure phosphatidic acid liposomes increased $\text{T}{}_{\mathrm{<mtext>t</mtext>}}$ only 1.5°C. However, application of 100 atm helium pressure to dipalmitoyl phosphatidic acid lipsomes containing 100 mmol methoxyflurane/mol lipid almost completely antagonized the effect of the anesthetic. A nonlinear pressure effect is observed. In a previous study, a concentration of 60 mmol methoxyflurane/mol dipalmitoyl phosphatidylcholine depressed $\text{T}{}_{\mathrm{<mtext>t</mtext>}}$ only 1.5°C, exhibiting a linear pressure effect. The completely different behavior in the charged membrane is best explained by extrusion of the anesthetic from the lipid phase.     

The diffusion-concentration product of oxygen in lipid bilayers using the spin-label T1 method

A method is described to measure the oxygen diffusion-concentration product, $\text{D}{}_{\mathrm{<mtext>o</mtext>}}\text{[}\text{O}{}_2\text{]}$, at any locus that can be probed or labeled using nitroxide radicals. The method is based on the dependence of the spin-lattice relaxation time $\text{T}{}_1$ of the spin label on the bimolecular collision rate with oxygen. Strong Heisenberg exchange between spin label and oxygen contributes directly to $\text{T}{}_1$ of the spin label, while dipolar interactions are negligible. Both time-domain and continuous wave saturation methods for studying $\text{T}{}_1$ are considered. The method has been applied to phospholipid liposomes using fatty acid spin labels. A discontinuity in $\text{D}{}_{\mathrm{<mtext>o</mtext>}}\text{[}\text{O}{}_2\text{]}$ at the main phase transition was observed.     

Influence of duration of incubation on zooplankton respiration and excretion results

Respiration (O), ammonium (NH₄), phosphate (PO₄), total nitrogen (N_T) and phosphorus (P_T) excretions were measured on mixed zooplankton during 3-, 6-, 9-, 12-, 21-, and 24-h incubation periods at 20–23 C. The excretion rates of PO₄, N_T. and P_T decrease during a 21-h period, while rates of respiration and excretion of NH{IN4} are constant. The percentage of inorganic nitrogen excreted increases regularly from 3 h (30–40% of total nitrogen) to 21 h (70–80%) and it could be either due to a bacterial activity which was measured or to a decrease with time of organic nitrogen excreted because of starvation. $\text{O}\text{N}{}_{\mathrm{T}}$, $\text{O}\text{PO}{}_4$, $\text{O}\text{P}{}_{\mathrm{T}}$, and $\text{NH}{}_4\text{PO}{}_4$ ratios increase during the first 9 h of incubation; the percentage of inorganic phosphorus excreted is higher at the very beginning and then remains constant from 6 to 24 h. $\text{O}\text{NH}{}_4$ and $\text{N}{}_{\mathrm{T}}\text{P}{}_{\mathrm{T}}$ ratios are constant during a 24-h term, which makes them useful metabolic indexes.     

The influence of charge on phosphatidic acid bilayer membranes

A complete titration of phosphatidic acid bilayer membranes was possible for the first time by the introduction of a new anaologue, $\text{1,2-}\text{dihexadecyl}\text{-sn-}\text{glycerol-3-phosphoric}$ acid, which has the advantage of a high chemical stability at extreme pH values. The synthesis of this phosphatidic acid is described and the phase transition behaviour in aqueous dispersions is compared with that of three ester phosphatidic acids; $\text{1,2-}\text{dimyristoyl}\text{-sn-}\text{glycerol-3-phosphoric}$ acid, 1,3-dimyristoylglycerol-2-phosphoric acid and $\text{1,2-}\text{dipalmitoyl}\text{-sn-}\text{glycerol-3-phosphoric}$ acid.The phase transition temperatures ($\text{T}{}_{\mathrm{<mtext>t</mtext>}}$) of aqueous phosphatidic acid dispersions at different degrees of dissociation were measured using fluorescence spectroscopy and 90° light scattering. The $\text{T}{}_{\mathrm{<mtext>t</mtext>}}$ values are comparable to the melting points of the solid phosphatidic acids in the fully protonated states, but large differences exist for the charged states.The $\text{T}{}_{\mathrm{<mtext>t</mtext>}}$ vs. pH diagrams of the four phosphatidic acids are quite similar and of a characteristic shape. Increasing ionisation results in a maximum value for the transition temperatures at pH 3.5 ($\text{p}\text{K}{}_1$). The regions between the first and the second $\text{p}\text{K}$ of the phosphatidic acids are characterised by only small variations in the transition temperatures (extended plateau) in spite of the large changes occurring in the surface charge of the membranes. The slope of the plateau is very shallow with increasing ionisation. A further decrease in the H⁺ concentration results in an abrupt change of the transition temperature. The slope of the $\text{T}{}_{\mathrm{<mtext>t</mtext>}}$ vs. pH diagram beyond $\text{p}\text{K}{}_2$ becomes very steep. This is the     

Serum principal iodothyronines and the influence of cold exposure associated with shearing in the sheep

The effect of cold exposure caused by shearing on serum thyroid hormone (TH) concentrations in sheep kept at an ambient temperature of 8.5°C was studied. While the deep body temperature fell to the lowest level 4 h after shearing the concentration of triiodothyronine (T₃) increased to a peak value at that time. Thyroxine (T₄) and metabolically inactive reverse triiodothyronine (rT₃) levels reached their peak value after 24 h. The $\text{T}{}_3\text{T}{}_4$ ratio reached a maximum at about 4 h and $\text{rT}{}_3\text{T}{}_4$ and $\text{rT}{}_3\text{T}{}_3$ ratios rose to maximum values about 24 h after shearing. This sequence of events suggest a biphasic response to cold—an immediate secretion of TH from the thyroid gland, followed by adaptive alteration in T₃ and rT₃ generation in the extrathyroidal tissues.     

Involvement of thyroid gland in the development of migratory disposition in the redheaded bunting, Emberiza bruniceps

In the redheaded bunting Emberiza bruniceps, thyroidectomy inhibited premigratory fattening and nocturnal restlessness—two characteristics of avian migration—observed in caged birds during the premigratory period (March/April). Thyroxine (T₄) and triiodothyronine (T₃) administration in thyroidectomized birds stimulated locomotor activity and restored the loss in body weight. Annual variations in circulating thyroid hormone concentrations revealed a significant rise in $\text{T}{}_3\text{T}{}_4$ ratio prior to spring migration in both years studied. This increase in circulating $\text{T}{}_3\text{T}{}_4$ ratio may be associated with the development of migratory disposition in this bird. There was no increase in circulating $\text{T}{}_3\text{T}{}_4$ ratio prior to autumnal migration, however, plasma T₄ increased significantly. Different thyroidal mechanisms are most likely involved in spring and fall migratory periods. While T₃ remained low throughout, apart from the characteristic spring rise, high T₄ levels in E. bruniceps were associated with periods of reproduction and molting, the latter coinciding partly with autumnal migration.     

Binding of bovine cytochrome b5 to phosphatidycholine liposomes Characterization of the reconstituted lipid-protein vesicles

Cytochrome $\text{b}{}_5$ was extracted and purified from beef liver by a detergent method (cytochrome $\text{d-b}{}_5$). The hydrophilic moiety which carries the heme group (cytochrome $\text{t-b}{}_5$) was prepared by trypsin action upon pure cytochrome $\text{d-b}{}_5$.Single-shelled lecithin liposomes form complexes with cytochromes $\text{d-b}{}_5$ up to a molar ratio of one protein for 35 phospholipids. The lipid-protein complexes were isolated by gel filtration on Sepharose 4B. They are hollow vesicles in which [³H]-glucose can be trapped. Their diameter is greater than that of the initial liposomes.Cytochrome $\text{t-b}{}_5$ does not interact with the vesicles. These results show that the hydrophobic tail is necessary for the binding and that the hydrophilic part of the protein is located on the outer face of the vesicles. This asymmetry is also proved by the action of reducing agents.Experiments with saturated phosphatidylcholines show that the protein interacts with the lipids both below the transition temperature $\text{T}{}_{\mathrm{M}}$. i.e. when the aliphatic chains are in a crystalline state, and above $\text{T}{}_{\mathrm{M}}$, when the alipathic chain are in a fluid state.¹H NMR spectra show that even at the maximum cytochrome $\text{d-b}{}_5$ concentration the presence of the proteins does not markedly change the dynamics to the phospholipid molecules. An asymmetric single-shelled vesicle structure is proposed for the complex.     

Electrophoretic measurements about the relation between transition voltage and ζ-potential of biological membranes

The effects of inorganic cations, $\text{n-}\text{hexanol}$, saccharose and ²H₂O on the electrophoretic mobility and ζ-potential of membrane vesicles from nerve myelin were measured and the results compared with the corresponding effects of the same reagents on the transition voltage, $\text{V}{}_{\mathrm{<mtext>Tr</mtext>}}$, of the nerve axon membrane. Different cation concentrations and ²H₂O affect both potentials, the ζ-potential and $\text{V}{}_{\mathrm{<mtext>Tr</mtext>}}$, in a parallel way. Saccharose and $\text{n-}\text{hexanol}$, however, shift $\text{V}{}_{\mathrm{<mtext>Tr</mtext>}}$ but leave the electrophoretic mobility of the myelin vesicles unchanged. These results suggest that $\text{V}{}_{\mathrm{<mtext>Tr</mtext>}}$ shifts are not necessarily linked to changes in the membrane surface charge density but may also be caused by an interaction between the reagent and non-polar groups of the membrane interior.     

Purification and characterization of a glycoprotein,from normal human liver,which has the same molecular weight and chemical properties as plasma α1-antrypsin

An $\text{α}{}_1\text{-}\text{mantitrypsin-like}$ material has been purified to homogeneity from the soluble fraction of normal human liver by procedures adapted from those employed for plasma $\text{α}{}_1\text{-}\text{antrypsin}$. The liver material, in contrast to a previous report¹ has the same molecular weight as the corresponding normal plasma $\text{α}{}_1\text{-}\text{antrypsin}$. The subunit structure, immunoelectrophoretic and immunological properties of the liver glycoprotein are identical to those of normal plasma $\text{α}{}_1\text{-}\text{antrypsin}$. Amino acid and carbohydrate compositions of the liver material are similar to those of $\text{α}{}_1\text{-}\text{antrypsin}$ obtained from the plasma. The $\text{α}{}_1\text{-}\text{antrypsin-like}$ material has also been isolated and purified from the microsomal fraction of liver It has the same molecular weight and immunological properties as glycoprotein obtained from the cytosol. Although inhibitors of lysosmal proteases were added during the homogenization of the liver, the purified glycoprotein is devoid of trypsin-inhibitory capacity. The loss of inhibitory activity could be due to extensive cellular autolysis before autopsy.     

Differential polarized phase fluorometric studies of phospholipid bilayers under high hydrostatic pressure

Differential polarized phase fluorometry was used to quantify the rotational rate ($\text{R}$) and limiting anisotropy ($\text{r}{}_{\mathrm{\infty }}$) of the membrane probe diphenylhexatriene (DPH) in solvents and lipid vesicles exposed to hydrostatic pressures ranging from 1 bar to 2 kbar. These measurements reveal the effect of pressure on the phase-transition temperatures of the phosphatidylcholine vesicles, and the effects of pressure on order parameter of the acyl side-chain region of the membranes, the latter as indicated by $\text{r}{}_{\mathrm{\infty }}$. In addition to the well-known elevation of the transition temperature ($\text{T}{}_{\mathrm{<mtext>c</mtext>}}$) with pressure, our results demonstrate that increased pressure restores the order of the bilayers to that representative of temperatures below the transition temperature. We also found that solvents which allowed free isotropic rotation of DPH at 1 bar no longer allowed free rotation when sufficiently compressed; moreover, the apparent DPH rotational rate increased with $\text{r}{}_{\mathrm{\infty }}$. Pressure studies using both DPH and the charged DPH analogue, trimethylammonium DPH (TMA-DPH) indicated that the $\text{T}{}_{\mathrm{<mtext>c</mtext>}}$ of dipalmitoylphosphatidylcholine vesicles increased 23 K/kbar and an apparent volume change of 0.036 ml/mol lipid at the phase transition. Assuming, as has been proposed, that TMA-DPH is localized near the glycerol backbone region of the bilayers, these results indicate a similar temperature- and pressure-dependent phase transition in this region and the acyl side-chain region of the membrane.     

A general method for the direct isolation of the 3'-terminal fragments derived from transfer RNA molecules

A general procedure for the isolation of 3′-linked fragments derived from tRNA molecules is described. Purified N-2-naphthoxyacetylglycyl derivatives of the $\text{tRNA}{}_1{}^{\mathrm{Gly}}$ and $\text{tRNA}{}_2{}^{\mathrm{Gly}}$ of yeast were exhaustively digested with RNase T₁ and the 3′-linked fragments (bearing the derivative) were separated from other degradation products (lacking the derivative) by stepwise chromatography on BD-cellulose. Subsequent chromatographic resolution and base-composition analysis allowed tentative identification of the 3′-terminals of $\text{tRNA}{}_1{}^{\mathrm{Gly}}$ and $\text{tRNA}{}_2{}^{\mathrm{Gly}}$ as Gp(Cp,Ap)CpCpA and Gp(Cp,Cp,Up,Ap)CpCpA, respectively. The potential utility of this procedure for development of a novel approach to nucleic acid sequence analysis is discussed.     

A restatement of melittin-induced effects on the thermotropism of zwitterionic phospholipids

Perturbations induced by melittin on the thermotropism of dimyristoyl-, dipalmitoyl-, distearoylphosphatidylcholine and natural sphingomyelin are investigated and rationalized from data obtained by fluorescence polarization, differential scanning calorimetry and Raman spectroscopy. Depending on the technique and / or experimental conditions used, the observed effects differ at the same lipid to protein molar ratio, due to partial binding of melittin. The binding is more efficient for tetrameric than for monomeric melittin, but in both cases its affinity is weaker for phosphatidylcholine dispersions in the gel phase than for sonicated vesicles. For temperatures $\text{T ? T}{}_{\mathrm{<mtext>m</mtext>}}$ efficient binding occurs whatever the initial state of the lipids is. One can summarize the effects induced by melittin on the transition temperature as follows: (i) No upward shift is observed on synthetic phosphatidylcholines when lipid degradation is avoided. This is achieved by using highly purified melittin, phospholipase inhibitors, and / or non-hydrolysable lipids. (ii) Melittin monomer does not change $\text{T}{}_{\mathrm{<mtext>m</mtext>}}$. (iii) When melittin tetramer is stabilized, it decreases $\text{T}{}_{\mathrm{<mtext>m</mtext>}}$ by 10–15 deg. C. The transition broadens, and is finally abolished for $\text{R}{}_{\mathrm{<mtext>i</mtext>}}\text{? 2}$. Very similar results are found for natural sphingomyelin. Fluorescence polarization indicates similar changes in order and dynamics of the acyl chains for all lipid studied. For $\text{T ? T}{}_{\mathrm{<mtext>m</mtext>}}$, fluorescence and Raman show that melittin decreases the amount of CH₂ groups in ‘trans’ conformation and the intermolecular order of the chains. According to fluorescence data, there is an increase of the rigid-body orientational order at $\text{T ? T}{}_{\mathrm{<mtext>m</mtext>}}$, while from Raman the positional intermolecular order decreases without significant change in the CH₂ groups ‘trans’/‘gauche’ ratio.     

Proton nuclear magnetic resonance studies of the manganese (II) binding site of concanavalin A. Effect of saccharides on the solvent relaxation rates

The longitudinal relaxation rate ($\text{1}\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>p</mtext>}}$) of water protons was studied in solutions of Mn(II)-concanavalin A at a number of frequencies. These relaxation rates were lowered in the presence of a variety of saccharides which have affinities for concanavalin A which range over two orders of magnitude. A good correlation was found in which saccharides which bind tightly have the greatest effect and saccharides which bind weakly or not at all have little effect on the $\text{1}\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>p</mtext>}}$ values. The temperature dependence of the proton relaxation rates showed that the lowering of these rates in the presence of saccharides was most likely due to a change in the exchange rate of solvent interacting with protein-bound Mn(II), $\text{1}\text{T}{}_{\mathrm{<mtext>m</mtext>}}$.An analysis of the temperature and frequency dependence of the $\text{1}\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>p</mtext>}}$ and $\text{1}\text{T}{}_{\mathrm{<mtext>2</mtext><mtext>p</mtext>}}$ (transverse) solvent proton relaxation rates resulted in evaluation of a number of parameters for solvent water molecules interacting in the first coordination sphere of Mn(II) bound to concanavalin A. The ratio of the number of water molecules (q) to the Mn(II)-proton distance (r) obtained from a computer fit of the data over a limited temperature range is in accord with the findings of Koenig et al. ((1973) Proc. Nat. Acad. Sci.70, 475) and Meirovitch and Kalb ((1973) Biochim. Biophys. Acta303, 258). However, our studies of $\text{1}\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>p</mtext>}}$ and $\text{1}\text{T}{}_{\mathrm{<mtext>2</mtext><mtext>p</mtext>}}$ of water over a more extensive temperature range are best fit with the following conclusions: at low temperatures (<20 °C), the data are consistent with an outer-sphere relaxation process. At higher temperatures (> 30 °C), the water molecule in the inner coordination sphere of the bound Mn(II) begins exchanging more rapidly and contributes to the relaxation processes ($\text{1}\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>p</mtext>}}$ and $\text{1}\text{T}{}_{\mathrm{<mtext>2</mtext><mtext>p</mtext>}}$). The relaxation time of protons in the inner coordination shell, T_1M, contributes over the entire temperature range and produces a frequency dependence in the relaxivity data from 6 to 100 MH_z since the contributions to the correlation times are in the range 10^?9-10^?8 sec.     

Cellular aspects of tolerance. IV. Strain variations of tolerance induceability

The antibody response to RGG of 8-wk old mice of various strains was assessed in terms of half-lives ($\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$) of lightly iodinated rabbit gamma globulin (¹³¹I-RGG) elimination. $\text{T}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ was increased if the small aggregate content in ¹³¹I-RGG was reduced. The effect of aggregates was least in AKR, largest in Balb/c and SJL and intermediate in the majority of strains, typified by A mice. Differences between various strains, in the degree of tolerance to aggregate freed RGG (centrifuged at 123,000 g), were observed. The level of residual responsiveness was greatest in SJL mice. More profound tolerance could be induced with biofiltered RGG. Resistance of SJL mice to tolerance induction was also observed when human gamma globulin (HGG) and bovine serum albumin served as tolerogen. Tolerance to RGG and sheep red cells was induced in cyclophosphamide-treated SJL animals.     

Hydration of Escherichia coli lipids: Deuterium T1 relaxation time studies of phosphatidylglycerol,phosphatidylethanolamine and phosphatidylcholine

The hydration properties of Escherichia coli lipids (phosphatidylglycerol, phosphatidylethanolamine) and synthetic $\text{1,2-}\text{dioleoyl}\text{-sn-}\text{glycero-3-phosphocholine}$ in H₂O/²H₂O mixtures (9:1, v/v) were investigated with ²H-NMR. Comparison of the ²H₂O spin lattice relaxation time ($\text{T}{}_1$) as a function of the water content revealed a remarkable quantitative similarity of all three lipid-H₂O systems. Two distinct hydration regions could be discerned in the $\text{T}{}_1$ relaxation time profile. (1) A minimum of 11–16 water molecules was needed to form a primary hydration shell, characterized by an average relaxation time of $\text{T}{}_1\text{≈ 90}\text{ms}$. (2) Additional water was found to be in exchange with the primary hydration shell. The exchange process could be described in terms of a two-site exchange model, assuming rapid exchange between bulk water with $\text{T}{}_1\text{= 500}\text{ms}$ and hydration water with $\text{T}{}_1\text{= 80–120}\text{ms}$. Analysis of the linewidth and the residual quadrupole splitting (at low water content) confirmed the size of the primary hydration layer. However, each lipid-water system exhibited a somewhat different linewidth behavior, and a detailed molecular interpretation appeared to be preposterous.