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Background: The impact of obesity on trauma outcomes had been equivocal and the majority of these studies were conducted in the Western population. The aim of our study is to evaluate the impact of obesity on morbidity and mortality in Singaporean patients after trauma., Methods: A retrospective case control study of trauma patients admitted to a tertiary institution was performed. Patients admitted for both blunt and penetrating trauma who are aged 16 years and above were included. Propensity score matching (PSM) was performed to reduce selection bias., Results: A total of 5239 patients were reviewed (obese 648, non-obese 4591). Median age was 76 (IQR 63-83) years. Median BMI was 22.8 kg/m 2 (IQR 20.4-25.2). The overall median ISS was 10 (IQR 9-14). Median ISS was higher in non-obese patients, 10 (IQR 9-14) as compared to obese patients of ISS 9 (IQR 9-13), p = 0.003. Obese patients had more abdominal injuries, n = 27/648 (4.2%) than non-obese n = 114/4591 (2.5%), p = 0.013. Non-obese patients had more head injuries, n = 1698/4591 (37%) as compared to the obese n = 195/648 (30.1%), p = 0.001. In-hospital mortality was comparable in the matched cohort (obese n = 11/645 (1.7%), (non-obese, n = 39/1290 (3.0%), OR 0.56 [0.28-1.09], p = 0.085). ICU admission was shorter for the obese (n = 36/645 (5.6%) versus n = 108/1290 (8.4%), OR 0.65 [0.44-0.96], p = 0.027). Obese patients had higher incidence of venous thromboembolism (n = 16/645 (2.5%) versus n = 12/1290 (0.9%), OR 2.71 [1.27-5.76], p = 0.007)., Conclusion: Obese trauma patients had fewer head injuries, fewer ICU admissions but comparable mortality rates. Obesity was also associated with higher incidence of venous thromboembolism in the matched cohort., (© 2022 Royal Australasian College of Surgeons.)

Background: In difficult gallbladders, partial or subtotal cholecystectomy (SC) has been described as a reasonable procedure with safe outcomes. Our aim was to look at our data on SC with respect to safety, morbidity and long-term outcome., Methods: A retrospective analysis was performed for 3560 patients undergoing cholecystectomy from January 2010 to June 2016. For patients who underwent SC, demographics, intra-operative and follow-up details were analysed., Results: A total of 168 SC patients were included. 102 (60.7%) were male while 66 (39.3%) were female. The median age was 63 years (31-87). These patients were on follow-up for a median of 29 months (1.7-80). 153 were attempted laparoscopically and there were 25 (16.3%) patients which had open conversion. The rest of the 15 patients had open SC. Mean operative time 150 min (70-315) and average blood loss was 170 ml (50-1500). Median length of stay for these patients was 4 days (1-68). There were no common bile duct (CBD) injuries. We had 12 (7.1%) post-operative collections, 4 (2.4%) wound infections, 1 (0.6%) bile leak and 7 (4.2%) retained stones. Post-operative endoscopic retrograde cholangiopancreatography (ERCP) was performed on 4 (2.4%) patients with successful retrieval of CBD stones. One patient has spontaneous passage of CBD stone. The rest of the two patients with very small retained stones in remnant gallbladder were successfully managed conservatively. There was no 30-day or operation-related mortality. No patient required a second operation., Conclusions: SC is safe and feasible when encountering a difficult gallbladder.

Purpose: To identify predictors of arteriovenous fistula (AVF) patency in Asian patients with autogenous radio-cephalic arteriovenous fistula (RCAVF)., Methods: Retrospective review of 436 RCAVFs created between 2009 and 2013. Predictors of patency were identified with univariate and multivariate analysis. Kaplan-Meier survival analysis and log-rank test were used to calculate patency rates., Results: Overall secondary patency rate was 72% at 12 months, 69% at 24 months, 58% at 36 months, 57% at 48 months, 56% at 60 months and 54% at 72 months. Univariate analysis showed that factors which predict for patency include male gender (p = 0.003), good diabetic control (p = 0.025), aspirin use (p = 0.031), pre-dialysis status (p = 0.037), radial artery diameter (p = 0.029) and non-calcified radial arteries (p = 0.002). Age (p = 0.866), cephalic vein diameter (p = 0.630) and surgeon grade (p = 0.472) did not predict for primary AVF failure. Multivariate analysis revealed the male gender to be an independent predictor for patency (odds ratio 1.99, p = 0.01). Subset analysis showed a significantly larger average radial artery diameter of 2.3 mm amongst males, as compared to 1.9 mm amongst females (p = 0.001) and no statistical difference in the average cephalic vein diameter., Conclusions: Within our Asian study population, 12-month patency rate of RCAVF is 72%, 69% at 24 months, 58% at 36 months, 57% at 48 months, 56% at 60 months and 54% at 72 months. Male gender is an independent predictor for RCAVF patency. In females or patients with calcified radial arteries, a more proximal AVF should be considered.

Progranulin (GRN) mutations causing haploinsufficiency are a major cause of frontotemporal lobar degeneration (FTLD-TDP). Recent discoveries demonstrating sortilin (SORT1) is a neuronal receptor for PGRN endocytosis and a determinant of plasma PGRN levels portend the development of enhancers targeting the SORT1-PGRN axis. We demonstrate the preclinical efficacy of several approaches through which impairing PGRN's interaction with SORT1 restores extracellular PGRN levels. Our report is the first to demonstrate the efficacy of enhancing PGRN levels in iPSC neurons derived from frontotemporal dementia (FTD) patients with PGRN deficiency. We validate a small molecule preferentially increases extracellular PGRN by reducing SORT1 levels in various mammalian cell lines and patient-derived iPSC neurons and lymphocytes. We further demonstrate that SORT1 antagonists and a small-molecule binder of PGRN₅₈₈₋₅₉₃, residues critical for PGRN-SORT1 binding, inhibit SORT1-mediated PGRN endocytosis. Collectively, our data demonstrate that the SORT1-PGRN axis is a viable target for PGRN-based therapy, particularly in FTD-GRN patients.

Increasing evidence suggests that soluble aggregates of amyloid-β (Aβ) initiate the neurotoxicity that eventually leads to dementia in Alzheimer's disease. Knowledge on soluble aggregate structures will enhance our understanding of the relationship between structures and toxicities. Our group has reported a stable and homogeneous preparation of Aβ(1-42) oligomers that has been characterized by various biophysical techniques. Here, we have further analyzed this species by solid state nuclear magnetic resonance (NMR) spectroscopy and compared NMR results to similar observations on amyloid fibrils. NMR experiments on Aβ(1-42) oligomers reveal chemical shifts of labeled residues that are indicative of β-strand secondary structure. Results from two-dimensional dipolar-assisted rotational resonance experiments indicate proximities between I31 aliphatic and F19 aromatic carbons. An isotope dilution experiment further indicates that these contacts between F19 and I31 are intermolecular, contrary to models of Aβ oligomers proposed previously by others. For Aβ(1-42) fibrils, we observed similar NMR lineshapes and inter-side-chain contacts, indicating similar secondary and quaternary structures. The most prominent structural differences between Aβ(1-42) oligomers and fibrils were observed through measurements of intermolecular (13)C-(13)C dipolar couplings observed in PITHIRDS-CT experiments. PITHIRDS-CT data indicate that, unlike fibrils, oligomers are not characterized by in-register parallel β-sheets. Structural similarities and differences between Aβ(1-42) oligomers and fibrils suggest that folded β-strand peptide conformations form early in the course of self-assembly and that oligomers and fibrils differ primarily in schemes of intermolecular organization. Distinct intermolecular arrangements between Aβ(1-42) oligomers and fibrils may explain why this oligomeric state appears off-pathway for monomer self-assembly to fibrils., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The accumulation of plagues of β-amyloid (Aβ) peptides in the brain is a hallmark of Alzheimer's disease (AD). The redox-active Cu and Fe complexes of Aβ can cause damage to the neurons potentially via reactive oxygen species (ROS). The significant metal-mediated oxidative activity of CuAβ suggests that its presence can be chemically devastating regardless whether it is a cause or a result of AD. Flavonoids exhibit various benefits to human health, attributable to their metal-binding and antioxidation activities to certain extents. Despite broad interests and extensive studies of their metal-binding properties and anti/pro-oxidation activities, these properties and the mechanisms of the activities toward metal-centered oxidation reactions have not been fully revealed and concluded. We report herein distinctive antioxidation mechanisms between two flavonoid families toward the oxidation reactions by CuAβ(1-20), wherein the flavonols quercetin (Qr) and myricetin (Mr) competitively inhibit the oxidation of catechol by CuAβ(1-20) with K(i) of 11.2 and 32.6 μM, respectively, whereas the flavanols catechin (Ct) and epicatechin (Et) are substrates with k(cat) = 1.01 × 10(-2) and 1.55 × 10(-3) s(-1) and K(m) = 0.94 and 0.55 mM, respectively. Qr has a nearly 10-fold higher antioxidative efficacy than Ct against the oxidation activity of CuAβ, while Ct is effectively oxidized, which further decreases its antioxidant capacity. Similar inhibition patterns are observed toward oxidation of the catecholamine neurotransmitter dopamine by CuAβ(1-20). Metal ions and CuAβ bind Qr with a 1:1 ratio under our experimental conditions through the α-ketoenolate moiety as determined by the use of Co(2+) and Yb(3+) as paramagnetic NMR probes. Unlike flavanols, which are merely suicide antioxidative substrates, flavonols bind to the metal center and prevent metal-mediated redox reactions. We suggest flavonols may serve as leads for drug discovery and/or as agents toward preventing metal-mediated oxidative stress due to AD and other disorders. Moreover, CuAβ shows 8.6- and 4.2-fold higher kinetic regioselectivity in terms of k(cat) and k(cat)/K(m), respectively, toward the peroxidation of Ct than that of the enantiomer Et, suggesting potential development of metallo-catalysts in regioselective catalysis by the use of metallopeptides as templates.

Endogenous amyloid-β (Aβ) oligomeric aggregates have been proposed as toxic agents in Alzheimer's disease (AD). Knowledge of their structures not only may provide insight into the basis of their neurotoxicities but also may reveal new targets for therapeutic drugs and diagnostic tools. However, the low levels of these Aβ oligomers have impeded structural characterization. Evidence suggests that the endogenous oligomers are covalently modified in vivo. In this report, we demonstrate an established mass spectrometry (MS) methodology called precursor ion mapping (PIM) that potentially may be applied to endogenous oligomer characterization. First, we illustrate the use of this PIM technique with a synthetic Aβ(1-40) monomer sample that had been cross-linked with transglutaminase (TGase) and digested with pepsin. From PIM analysis of an Aβ(4-13) MS/MS fragment, precursor ions were identified that corresponded to peptic fragments of three TGase cross-linked species: Aβ(4-19)--(4-19), Aβ(4-19)--(20-34), and Aβ(1-19)--(20-34). Next, we demonstrate the applicability of the PIM technique to an endogenous Aβ sample that had been purified and concentrated by immunoaffinity chromatography. Without pepsin digestion, we successfully identified the full length and C-terminally truncated monomeric Aβ species 1-35 to 1-42, along with select methionine-oxidized counterparts. Because PIM focuses only on a subpopulation of ions, namely the related precursor ions, the resulting spectra are of increased specificity and sensitivity. Therefore, this methodology shows great promise for structural analysis and identification of post-translational modification(s) in endogenous Aβ oligomers.

The peptidyl antibiotic bacitracin (Bc) is one of the most widely used antibiotics which can bind divalent transition metal ions, including Mn(II), Co(II), Ni(II), Cu(II), and Zn(II). The metal binding is essential for its antimicrobial activity. Previous analysis of the hyperfine-shifted (1)H NMR signals of Co(II)-Bc A(1) revealed the structure of the metal binding environment and a potential hydrophobic site important for the bioactivity of this antibiotic. Co(II)-Bc in DMSO shows relatively sharper hyperfine-shifted (1)H NMR signals compared with the spectrum acquired in an aqueous solution, allowing more thorough analysis of the signals with 1D and 2D NMR methods. Pyrophosphate and derivatives bind to Co(II)-Bc to form kinetically inert ternary complexes. The coordinated D-Glu-4 is found detached from the metal center of metallobacitracin upon trimetaphosphate binding, implying its role in the antibiotic activity of Bc. We further demonstrate in this report the structure-function relationship on desamido-Bc of low antibiotic activity by the use of NMR, wherein D-Glu-4 is suggested to be important for the bioactivity of Bc. The interaction of the phospho-moiety with Bc is also reflected by DNA binding, wherein metal-free Bc does not bind DNA, whereas various metal complexes of Bc do. Cu(II)-Bc was further demonstrated to bind and oxidatively cleave DNA under reduction conditions in the air. It also exhibited a significant oxidative activity toward catechol oxidation, showing enzyme-like saturation kinetics with k(cat) = 7.0 x 10(-3) s(-1) and k(cat)/K(m) = 2.1 M(-1) s(-1) aerobically and k(cat) = 0.38 s(-1) and k(cat)/K(m) = 14.7 M(-1) s(-1) in the presence of 32 mM of H(2)O(2). The binding of pyrophosphate moiety to metallobacitracin, the detachment of d-Glu-4, and the significant oxidative activity of Cu(II)-Bc provide further insights into the bioactivity of this metallopeptide and Cu-oxygen chemistry.

The dinuclear aminopeptidase from Streptomyces griseus (SgAP) and its metal derivatives catalyze the hydrolysis of the phosphoester bis(p-nitrophenyl) phosphate (BNPP) and the phosphonate ester p-nitrophenyl phenylphosphonate with extraordinary rate enhancements at pH 7.0 and 25 degrees C [A. Ercan, H. I. Park, L.-J. Ming, Biochemistry 45, (2006) 13779-13793.], reaching 6.7 billion-fold in terms of the first-order rate constant of the di-Co(II) derivative with respect to the autohydrolytic rates. Since phosphoesters are transition state-like inhibitors in peptide hydrolysis, their hydrolysis by SgAP is quite novel. Herein, we report the investigation of this proficient alternative catalysis of SgAP and the role of each metal ion in the dinuclear site toward peptide and BNPP hydrolysis. Mn(II) selectively binds to one of the dinuclear metal sites (M1), affording MnE-SgAP with an empty (E) second site for the binding of another metal (M2), including Mn(II), Co(II), Ni(II), Zn(II), and Cd(II). Peptide hydrolysis is controlled by M2, wherein the k(cat) values for the derivatives MnM2-SgAP are different yet similar between MnCo- and CoCo-SgAP and pairs of other metal derivatives. On the other hand, BNPP hydrolysis is affected by metals in both sites. Thus, the two hydrolytic catalyses must follow different mechanisms. Based on crystal structures, docking, and the results presented herein, the M1 site is close to the hydrophobic specific site and the M2 site is next to Tyr246 that is H-bonded to a coordinated nucleophilic water molecule in peptide hydrolysis; whereas a coordinated water molecule on M1 becomes available as the nucleophile in phosphodiester hydrolysis.

A neuropathological hallmark of Alzheimer's disease (AD) is the presence of large numbers of senile plaques in the brain. These deposits are rich in fibrils that are composed of 40- and 42-residue amyloid-beta (Abeta) peptides. Several lines of evidence indicate that soluble Abeta aggregates as well as fibrils are important in the etiology of AD. Low levels of endogenous soluble Abeta aggregates make them difficult to characterize, but several species in extracts of AD brains have been detected by gel electrophoresis in sodium dodecyl sulfate (SDS) and immunoblotting. Individual Abeta oligomers ranging in size from dimers through dodecamers of 4 kDa monomeric Abeta have been resolved in other laboratories as discrete species by size exclusion chromatography (SEC). In an effort to reconstitute soluble Abeta aggregates in vitro that resemble the endogenous soluble Abeta aggregates, we previously found that monomeric Abeta(1-42) rapidly forms soluble oligomers in the presence of dilute SDS micelles. Here we extend this work in two directions. First, we contrast the size and secondary structure of these oligomers with those of synthetic Abeta(1-42) fibrils. SEC and multiangle light scattering were used to obtain a molecular mass of 150 kDa for the isolated oligomers. The oligomers partially dissociated to monomers through nonamers when incubated with SDS, but in contrast to endogenous oligomers, we saw no evidence of these discrete species prior to SDS treatment. One hypothesis to explain this difference is that endogenous oligomers are stabilized by covalent cross-linking induced by unknown cellular agents. To explore this hypothesis, optimal mass spectrometry (MS) analysis procedures need to be developed for Abeta cross-linked in vitro. In our second series of studies, we began this process by treating monomeric and aggregated Abeta(1-42) with three cross-linking agents: transglutaminase, glutaraldehyde, and Cu(II) with peroxide. We compared the efficiency of covalent cross-linking with these agents, the effect of cross-linking on peptide secondary structure, the stability of the cross-linked structures to thermal unfolding, and the sites of peptide cross-linking obtained from proteolysis and MS.

Histatin-5 (Hn5) is an antimicrobial salivary peptide of 24 amino acids. Two specific metal-binding sites were revealed with electronic, NMR, and EPR spectroscopy. The complex Cu(2)(II)-Hn5 effectively oxidizes catechol, exhibiting enzyme-like kinetics (k(cat)=0.011 and 0.060 s(-1) and k(cat)/K(m)=19 and 50 M(-1)s(-1) without and with 12.8mM H(2)O(2), respectively). The significant oxidative activity may contribute to the biological activity of this antibiotic metallopeptide.

The Cu2+ complexes of the 1-16 and the 1-20 fragments of the Alzheimer's disease-related beta-amyloid peptide (CuAbeta) show significant oxidative activities toward a catechol-like substrate trihydroxylbenzene and plasmid DNA cleavage. The latter reflects possible oxidative stress to biological macromolecules, yielding supporting data to the pathological role of these soluble Abeta fragments. The former exhibits enzyme-like kinetics and is dependent on [H2O2], exhibiting k(cat) of 0.066 s-1 (6000-fold higher than the reaction without CuAbeta) and k(cat)/Km of 37.2 m-1s-1 under saturating [H2O2] of approximately 0.24%. This kinetic profile is consistent with metal-centered redox chemistry for the action of CuAbeta. A mechanism is proposed by the use of the catalytic cycle of dinuclear catechol oxidase as a working model. Trihydroxylbenzene is also oxidized by CuAbeta aerobically without H2O2, affording rate constants of 6.50x10(-3) s-1 and 3.25 m-1s-1. This activity is also consistent with catechol oxidase action in the absence of H2O2, wherein the substrate binds and reduces the Cu2+ center first, followed by O2 binding to afford the mu-eta2:eta2-peroxo intermediate, which oxidizes a second substrate to complete the catalytic cycle. A tetragonally distorted octahedral metal coordination sphere with three coordinated His side chains and some specific H-bonding interactions is concluded from the electronic spectrum of CuAbeta, hyperfine-shifted 1H NMR spectrum of CoAbeta, and molecular mechanics calculations. The results presented here are expected to add further insight into the chemistry of metallo-Abeta, which may assist better understanding of the neuropathology of Alzheimer's disease.

In this study, a methacrylate-based orthopaedic bone cement was modified for use as a root canal sealer by changing the monomer from n-butyl methacrylate to tetrahydrofurfuryl methacrylate. The sealing and bonding capabilities, solubility and handling properties of the resin were evaluated. Seventy-three extracted mature human teeth with single canals were used. The teeth were randomly divided into two experimental groups each of 30 teeth; three control teeth were set aside for the leakage tests. Ten teeth were reserved for the scanning electron microscope studies. All the experimental teeth were root filled with gutta-percha and the resin sealer. The particle size of the polymer was limited in Group 1 to 75 microns or less and in Group 2 to 45 microns or less. The controls were filled with gutta-percha only. Silver nitrate solution was used as the leakage indicator. In Group 1 the mean linear penetration of the indicator was 1.10 mm +/- 1.04; four roots showed no leakage. In Group 2 the mean linear penetration of the indicator was 0.57 mm +/- 0.65; 12 roots showed no leakage. The test bodies showed no weight change after 1 and 3 months' storage in saline at 37 degrees C. The scanning electron microscope examination revealed a bond both between the resin and the dentine and between the resin and the gutta-percha. The working time of the new resin composition was normally about 50 min, and handling was found to be easy.

It has been claimed that glass-ionomer cements possess properties that could make them the "ideal' restorative material. In the light of current disquiet about the safety of dental amalgam, are glass-ionomer cements likely to become the replacement materials of choice?

This study compared the linear leakage patterns shown by 5% aqueous solution of methylene blue (MB) and India ink (II). One-hundred and twenty-five roots with single canals from extracted human teeth were used. The canals were cleaned and shaped using the step-back technique under copious irrigation. Three groups were arranged to receive root fillings of gutta-percha and one of three proprietary sealers using standard lateral condensation techniques: Sealapex (40 roots), Tubli-Seal (40 roots) and Ketac-Endo (20 roots). The remaining roots were used as controls: positive with gutta-percha and no sealer and negative with the apices sealed with nail varnish. The roots were randomly selected for 7-day immersion in either MB or II, then sectioned longitudinally; dye penetration was assessed using a Wild Leitz stereomicroscope. In all groups MB showed more penetration than II. The results were statistically significant at the P = 0.01 level, with the exception of the positive control groups. Negative controls showed no penetration. It is concluded that MB, which has a low molecular weight penetrated more deeply along root canal fillings than II, which has a large particle size.

Various surface treatments affected the shear bond strength of a dual-cured resin cement to a post-cured composite. Disks of post-cured composite inlay 2.0 mm thick by 10.0 mm in diameter, were mounted in acrylic cylinders. Various surface treatments were tested including: 1) smooth material; 2) treated by sandblasting (air abrasion); 3) sandblasting and etching with hydrofluoric acid (HF); 4) sandblasting and silane priming; 5) sandblasting, HF etching and silane priming; 6) HF etching alone; 7) HF etching and silane priming; and 8) silane priming alone. Specimens of dual-cured cement, 4.0 mm in diameter, were light-cured onto the prepared surfaces. The samples were stored in water at 37 degrees C for 7d before shear bond testing. The highest mean shear bond strengths were achieved for specimens with the following surface treatments: 1) by a combination of sandblasting, HF etching and silane priming; 2) sandblasting and HF etching; and 3) sandblasting and silane priming. Most bond failures in these groups were cohesive within the post-cured composite inlay disks. The bond strengths of the specimen in other groups were significantly lower and bond failures were mainly adhesive at the interface between the disk of post-cured composite and dual-cured resin cement.

The deformation of a cylinder in the diametral mode proved to be a linear function of the applied load for silicate cements, glass ionomer cements and glass. Two theoretical analyses were investigated, namely, Hertzian indenter theory and integration of the Hondros diametral theory. Whilst both theories predicted the experimentally observed linear behaviour, neither were quantitatively correct. Furthermore, the Hondros theory contained an experimentally inaccessible parameter, the angle subtended by the contact width at the centre.

Glass-ionomer cements have unique and varied properties: they are tough, translucent, biologically inert, resistant to degradation in the mouth, adhesive to tooth structure and they inhibit caries. In a number of ways they are probably the one substance available today that most approaches the ideal restorative material. In Part 1 of this two-part article the authors reviewed the physical and clinical properties of glass-ionomer cements, here they go on to describe their use in practical and sometimes novel restorative procedures.

Two recent proprietary glass ionomer cements showed that in the early stages of set the changes in relative permittivity (dielectric constant) and resistivity were rapid. However, when set, measurements indicated that the cements were highly ionic and polar and behaved similarly to the glass ionomer cements previously reported.

The unique properties of the glass-ionomer (or polyalkenoate) cement have widened the compass of restorative and preventive dentistry and changed traditional practice methods. Glass-ionomer cements are the reaction product of an aluminosilicate glass and a polyacid, for example poly (acrylic acid).

Thermal diffusivity, a property related to the thermal insulative efficiency of a material, was measured in nine glass-ionomer cements and compared with results from a silicate and a polycarboxylate cement. Each cement was mixed at various powder-liquid ratios (P/L) and moulded into a rectangular prism of approximate dimensions 2 cm cube with a thermocouple embedded in it. The prism was immersed in a constant-temperature bath at 1 degree C, and the fall in temperature was observed over a period of three min. Except for the initial and later stages, the plot of the logarithmic difference between external and internal temperatures of each block of cement against time showed a straight line in accord with theoretical prediction. From the slope, the thermal diffusivity of the material was calculated. The values for the silicate, polycarboxylate, and glass-ionomer-metal (cermet) showed a marked rise with increasing P/L, whereas at higher P/L, glass ionomer cements showed gradual change, with values being only slightly higher than the thermal diffusivity of dentin. Glass-ionomer cements are good thermal insulators over a wide range of P/L, and close agreement between experimental and theoretical data shows that glass-ionomer cements are homogenous isotropic materials.

Long-term elution of fluoride ions has been studied for one silicate and six glass-ionomer cements over a period of some two and-a-half years. Because equilibrium was not reached, classical diffusion theory could not be applied to the process. It was postulated that two elution processes were taking place, one short-term and rapid, the other more gradual and prolonged. It was possible only to estimate the maximum value for the diffusion coefficients.