Prevent Gel Formation Research Articles

Impact of Alternative Thermal and Non-Thermal Processing on the Enzymatic Activity of Papaya and Strawberry Nectars and Their Blends

Pectin methylesterase (PME) in papaya nectar results in undesirable gel formation and peroxidase (POD) in strawberry nectar leads to nutrient loss, browning, and off-flavor production. Because of this, the effect of alternative processing techniques including ultra high temperature (UHT, 20-135°C, 1-3 s), high pressure processing (HPP, 20 or 60°C, 200-600 MPa) and irradiation (0-10 kGy) on PME and POD activity in papaya and strawberry nectar and their respective blends were compared to traditional thermal processing (80-130°C, 0-10 min). Traditional thermal (110°C, 5 min, 71.5% reduction) and UHT (110°C, 1-3 s, 98.0% reduction) processing were able to sufficiently reduce PME activity and prevent gel formation in papaya nectar. PME reduction was enhanced by synergistic reductions in nectar blends after UHT at 80°C. HPP was unable to prevent gel formation in papaya nectar, with enhanced activity at 400 MPa. Exposure of a blend 50P:50S to 10 kGy irradiation prevented gel formation. UHT enhanced POD activity at 110°C and synergistic reductions resulted for POD activity in nectar blends after irradiation. These findings highlight the benefits of alternative processing in reducing enzyme activity in fruit nectars and nectar blends.

Human platelet lysate efficiency, stability, and optimal heparin concentration required in culture of mammalian cells.

BackgroundFetal bovine serum (FBS) has been used to support the growth and proliferation of mammalian cells for decades. Owing to several risk factors associated with FBS, several trials have been conducted to evaluate substitutes to FBS with the same efficiency and the lower risk issues.MethodsIn this study, human platelet lysate (HPL) derived from activated human platelets was evaluated as an alternative to FBS due to the associated risk factors. To evaluate the efficiency of the preparation process, platelet count was performed before and after activation. The concentrations of several growth factors and proteins were measured to investigate HPL efficiency. HPL stability was studied at regular intervals, and optimal heparin concentration required to prevent gel formation in various media was determined. The biological activity of HPL and FBS was compared by evaluating the growth performance of Vero and Hep-2 cell lines.ResultsResult of platelet count assay revealed the efficiency of HPL preparation process. Growth factor concentrations in HPL were significantly higher than those in FBS, while the protein content of HPL was lower than that of FBS. Stability study data showed that the prepared HPL was stable for up to 15 months at −20℃. Ideal heparin concentration to be used in different media was dependent on calcium concentration. Results of cell viability assay showed that HPL was superior to FBS in supporting the growth and proliferation of Vero and Hep-2 cells.ConclusionThe HPL prepared by the mechanical activation of platelets may serve as an efficient alternative to FBS in cell culture process.

Rheological behavior of suspensions of modified and unmodified cellulose nanocrystals in dimethyl sulfoxide

The rheological behavior of cellulose nanocrystal (CNC) and modified CNC (mCNC) suspensions in dimethyl sulfoxide (DMSO) was investigated. The efficiency of the surface modification of CNCs by grafting an organic acid chloride to produce hydrophobic CNCs has been verified by X-ray photoelectron spectroscopy (XPS). The thermal degradation temperature of the mCNCs was found to be 165 versus 275 °C for CNCs. The CNC suspensions in DMSO at 70 °C underwent gelation at very low concentration (1 wt%) after 1 day. The network formation was temperature sensitive and did not occur at room temperature. For gels containing 3 wt% CNCs, the complex viscosity at 70 °C increased by almost four decades after 1 day. For the mCNCs in DMSO, a weak gel was formed from the first day and temperature did not affect the gelation. Finally, the effect of adding 10 wt% of polylactide (PLA) to the solvent on the rheological properties of CNC and mCNC suspensions was investigated. The properties of suspensions containing 1.9 wt% CNCs and mCNCs increased during the first and second days, and PLA did not prevent gel formation. However, the reduced viscosity and storage modulus of the CNC and mCNC gels with PLA were lower than those of samples without PLA.

Viscosity-Reducing Bulky-Salt Excipients Prevent Gelation of Protein, but Not Carbohydrate, Solutions.

The problem of gelation of concentrated protein solutions, which poses challenges for both downstream protein processing and liquid formulations of pharmaceutical proteins, is addressed herein by employing previously discovered viscosity-lowering bulky salts. Procainamide-HCl and the salt of camphor-10-sulfonic acid with L-arginine (CSA-Arg) greatly retard gelation upon heating and subsequent cooling of the model proteins gelatin and casein in water: Whereas in the absence of additives the proteins form aqueous gels within several hours at room temperature, procainamide-HCl for both proteins and also CSA-Arg for casein prevent gel formation for months under the same conditions. The inhibition of gelation by CSA-Arg stems exclusively from the CSA moiety: CSA-Na was as effective as CSA-Arg, while Arg-HCl was marginally or not effective. The tested bulky salts did not inhibit (and indeed accelerated) temperature-induced gel formation in aqueous solutions of all examined carbohydrates-starch, agarose, alginate, gellan gum, and carrageenan.

Electrically released iron for fouling control in membrane bioreactors: A double-edged sword?

Iron injection through electrolytically mediated oxidative dissolution of an iron electrode has been proposed as a convenient means of inducing coagulation of wastewater contaminants and thereby reducing the extent of membrane fouling in membrane bioreactors (MBRs). In order to investigate the effect of electrically released iron on membrane fouling propensity, alginate was selected as a model soluble microbial product, and the impact of electrolyzed iron on the specific resistance to filtration (SRF) quantified for a range of electrolysis times. It was confirmed that ferrous iron (Fe(II)) was initially released with concentration increasing proportionally with time of application of electric current in agreement with Faraday's Law. The fouling propensity of alginate increased at early loading times due to the formation of alginate gels and highly stabilized colloidal species. The presence of calcium ions was found to exacerbate this gelation process as a result of i) alginate bridging by Ca2+ ions, and ii) the reduced capacity of the alginate to bind iron and stabilize iron oxide particles. Significant reduction in fouling propensity was observed at higher iron loadings as a result of the formation of substantive amounts of iron oxides which, most likely, adsorbed alginate and prevented gel formation.

Heparin concentration is critical for cell culture with human platelet lysate

Background aimsCulture media for mesenchymal stromal cells (MSCs) are generally supplemented with fetal bovine serum. Human platelet lysate (hPL) has been proven to be a very effective alternative without the risk of xenogeneic infections or immune reactions. In contrast to fetal bovine serum, hPL comprises plasma, and anticoagulants—usually unfractionated heparin (UFH)—need to be added to prevent gel formation. MethodsCultures of MSCs in hPL media with various concentrations of UFH and enoxaparin, a low-molecular-weight heparin (LMWH), were systematically compared with regard to proliferation, fibroblastoid colony-forming unit frequency, immunophenotype and in vitro differentiation. ResultsAt least 0.61 IU/mL UFH or 0.024 mg/mL LMWH was necessary for reliable prevention of coagulation of hPL pools used in this study. Higher concentrations impaired cellular proliferation in a dose-dependent manner even without benzyl alcohol, which is commonly added to heparins as a bacteriostatic agent. Colony-forming unit frequency was also reduced at higher heparin concentrations, particularly with LMWH, whereas no significant effect was observed on cellular morphology or immunophenotype. High concentrations of heparins reduced the in vitro differentiation toward adipogenic and osteogenic lineages. ConclusionsHeparin concentration is critical for culture of MSCs in hPL media; this is of particular relevance for cellular therapy where cell culture procedures need to be optimized and standardized.

Matrix Metalloproteinase-8 Is Involved in Dermal Nerve Growth: Implications for Possible Application to Pruritus from In Vitro Models

Cutaneous nerve density is related to abnormal itch perception in dermatoses, such as atopic dermatitis and xerosis. However, the mechanisms underlying the elongation of dermal nerve fibers within the interstitial collagen (CoL) matrix are poorly understood. In this study, a culture system of rat dorsal root ganglion neurons consisting of type I CoL and a Boyden chamber containing a nerve growth factor (NGF) concentration gradient was used. Nerve fibers penetrating into type I CoL gel were observed in the presence of the NGF concentration gradient. Levels of matrix metalloproteinase-8 (MMP-8) mRNA and protein were increased in the cultured neurons and the conditioned medium, respectively. The nerve fiber penetration was dose dependently inhibited by MMP-8 blockers. Moreover, MMP-8 immunoreactivity was partially localized at growth cones in NGF-responsive nerve fibers. Semaphorin 3A stimulation also showed the opposite effects on these NGF-dependent events. Intriguingly, MMP-8 expression was upregulated by type I and III CoLs, which are substrates for this enzyme. These results suggested that MMP-8 is involved in sensory nerve growth within the interstitial CoL matrix through modulation by the axonal guidance molecules and/or extracellular matrix components. These findings provide insight into the development of pruritus involving skin nerve density.

Direct formation of “artificial wool” nanofiber via two‐spinneret electrospinning

AbstractTo reproduce the excellent characteristics of natural fibers like wool and proteins, a novel two‐spinneret electrospinning technology was demonstrated in this communication, which can generate three‐dimensional self‐crimp fibers of HSPET/PTT, HSPET/PAN and PU/PAN directly. In the apparatus, two spinnerets were used to prevent gel formation or precipitation of the polymer, and the voltages of opposite polarities were applied to the spinnerets respectively. And the elecctrospun fibers morphology was observed by using scanning electron microscopy (SEM). © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2012

Copper-mediated ATRP of MMA in DMSO from unprotected dextran macroinitiators

To easily perform atom transfer radical polymerization (ATRP) of methyl methacrylate (MMA) in homogeneous medium from unprotected polysaccharidic macroinitiators, the ATRP of MMA carried out in dimethylsulfoxide has been firstly reinvestigated using Cu(I)Br as metal catalyst and ethyl 2-bromoisobutyrate as initiator. Two ligands, 2,2′-bipyridyne (Bpy) and N-(n-propyl)-2-pyridylmethanimine (n-Pr-PMI), have been compared, as well as the addition of Cu(II)Br2 (10 molar % relative to Cu(I)Br) or the adjustment of experimental temperature (from 30 to 60 °C). Appropriate conditions were founded for both ligands to achieve a controlled ATRP. Bpy giving far much faster polymerization compared to n-Pr-PMI, n-Pr-MI was preferred as ligand for accurate tuning of the conversion. Experimental procedure was then applied to unprotected macroinitiators based on dextran to obtain amphiphilic poly(methyl methacrylate) grafted dextran. While homopolymerization was controlled in these conditions, parameters have to be adjusted for the hydrophilic unprotected macroinitiators to prevent gel formation and control ATRP.

Reversible Heat-Set Organogel Based on Supramolecular Interactions of β-Cyclodextrin in N,N-Dimethylformamide

This paper describes the first reversible, heat-set organogel based on the supramolecular interactions of beta-cyclodextrin (beta-CD). The gel was prepared by interaction of diphenylamine (DPA) with beta-CD and lithium chloride in N,N-dimethylformamide (DMF). In this gel system, DPA could be gelated in DMF as the temperature increased and then dissolved again as the temperature decreased. In the microscopic structure of gel, beta-CDs play a key role in the formation of nanorods and microfibers. Some important features of the gel were observed. (1) The system is a multicomponent solution, in which each of the four components is required for the organogelation property. (2) The system is a reversible, thermo-responsive organogel composed of small organic molecules. When the temperature is lower than T(gel), the gel transforms back into a solution. The reversible thermo-transition was confirmed by differential scanning calorimetry (DSC). The gel system is responsive to the concentration of LiCl. No gel was formed without LiCl. The stimuli responses of the system with other salts such as KCl and NaCl were weaker than with LiCl. (4) The system is responsive to the addition of guest molecules. The structures and sizes of the guest molecules could influence the gel formation. Generally, T(gel) decreased by adding guest molecules in the gel system, but some guest molecules, whose structures are exactly fitted to the cavity of CDs, could prevent gel formation. This work may provide new avenues in delivery of functional molecules as well as design of intelligent materials and biomaterials.

Comparative study of the effect of sulfur on the morphology and rheological properties of SB‐ and SBS‐modified asphalt

AbstractThe modification of asphalt with styrene‐ butadiene block copolymers and sulfur was studied to elucidate the effect of the molecular characteristics of the polymer, polymer content, and sulfur/polymer ratio on the physical properties of modified asphalts. Two types of styrene‐butadiene copolymers were used (SB and SBS), which differed considerably in terms of their chain architecture, average molecular weights, and size and distribution of their polybutadiene and polystyrene blocks, as shown by gel permeation chromatography, infrared spectroscopy, nuclear magnetic resonance, and differential scanning calorimetry. Sulfur/polymer/asphalt blends were prepared by a hot mixing process and characterized by conventional tests, fluorescence microscopy, and rheology. The results revealed that the morphology of the blends is strongly dependent on polymer concentration and sulfur/polymer ratio. In‐depth rheological characterization showed that the thermomechanical properties changed considerably upon addition of small amounts of sulfur. Collectively, these results suggest that sulfur increases the compatibility between polymer and asphalt by crosslinking polymer chains. Interestingly, the rheological behavior of blends prepared with a combination of SB and sulfur was similar to that exhibited by blends prepared with SBS either in the presence or absence of sulfur. This is explained by assuming that the addition of small amounts of sulfur to SB‐modified asphalt facilitates the formation of an elastomeric network that resembles the one found in SBS‐modified asphalt, effectively contributing to asphalt reinforcement. Nonetheless, the exact dosage of sulfur must be carefully controlled to prevent gel formation. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2010

Effect of Branching on Polymer Diffusion in Branched Poly(butyl methacrylate) Latex Films

Fluorescent donor- and acceptor- labeled latex particles comprised of branched poly(butyl methacrylate) (PBMA) were prepared via semicontinuous emulsion polymerization. All latex particles have similar particle size and narrow size distribution. The degrees of branching of the PBMA samples were controlled by adding various amount of bisphenol A dimethacrylate (BPDM) as a branching agent. 1-Dodecanethiol (C12−SH) was used as a chain transfer agent to prevent gel formation and control the molecular weight. The number average molecular weights of all PBMA samples are ca. 45 000 g/mol. The degrees of branching were determined by 1H NMR. Latex films were cast from a 1:1 mixture of donor- and acceptor-labeled latices. The polymer diffusion in the latex films was studied by measuring the energy transfer (ET) from the donor to the acceptor using the time-correlated single photon counting technique. Polymer diffusion rates increased with decreasing values of Tg when the latex films were annealed at 23 °C. After co...

Improved method for the determination of biogenic amines and polyamines in vegetable products by ion-pair high-performance liquid chromatography

Here, we report on the optimisation and validation of a liquid chromatographic method for the determination of 12 biologically active amines from vegetal food products in a single 40-min run. The suitability of the method was checked in five vegetal products of distinct matrix: spinach (leaves), hazelnut (high protein and fat content), banana, potato (high starch content), and milk chocolate (processed). Sample preparation consisted of a 0.6 M perchloric acid extraction from a minced homogeneous aliquot. For samples with high starch content, a previous mild hydrolytic treatment was required to prevent gel formation. The range of linearity was from 0.1 to 10 mg/l, except for serotonin and spermine (from 0.5 to 10 mg/l), and the correlation coefficient was higher than 0.997 ( P < 0.001) for all standard curves. The detection limits and the determination limit were below 0.07 and 0.2 mg/l, respectively, except for spermine, which was 0.14 and 0.4 mg/l. The precision of the method was satisfactory; the relative standard deviation obtained for each amine in each product was acceptable according to Horwitz. Recovery was between 77 and 110% for all amines, irrespective of the product.

Use of Carbodiimides as Stabilizing Agents to Deliver Water—Labile Active Ingredients in Liquid Systems Including Aqueous Medium—Amitraz as a Case Study

Abstract Several active ingredients such as sulfonylureas and diimidines (Amitraz) are unstable in water. They undergo hydrolysis producing metabolites that are no longer biologically active. The rate of hydrolysis is a function of several factors, such as the solvent system, access to water, pH, temperature, ionic strength, and the types of emulsifiers used in the liquid matrices. Some of the active ingredients undergo rapid hydrolysis even with the residual water present in the emulsifier system in an organic emulsifiable concentrate formulation. It is therefore difficult to formulate such active ingredients in liquid delivery systems. Use of carbodiimides like Stabaxol I® (bis-2,6-diisopropylphenylcarbodiimide) as a water scavenger to stabilize oil-based formulations for Amitraz is well documented and is commercially practiced. We have found that unique combinations of carbodiimides with specific solvent-emulsifier-polymer blends in a buffered system can produce stable microemulsions of Amitraz in water. Such aqueous formulations with 200–500 ppm Amitraz in &amp;gt;99 % water, were stable with 95 % retention on accelerated storage at 50°C for two weeks. This data translates to greater than two years stability at room temperature. Such formulations may be extended to deliver sulfonylureas. We also observed that the concentrate and the matrix containing Stabaxol I® and emulsifiers can produce gels on storage, adversely affecting physical stability. We have identified several additives and alternatives to prevent gel formation enabling long-term storage stability of the matrix and the concentrate. Examples with supporting data will be provided. A rationale and proposed mechanism for the enhanced stability and potential biological activity will be discussed.

Hydrogenation and neutralization of carboxylic styrene–butadiene latex to form thermoplastic elastomer with excellent thermooxidation resistance

AbstractHydrogenation of carboxylic styrene–butadiene rubber latex was carried out using hydrazine and hydrogen peroxide with ferrous sulfate as a catalyst without pressurized hydrogen and an organic solvent. A mixed inhibitor was used during hydrogenation to prevent gel formation. Various hydrogenation conditions were studied. Ferrous sulfate is better than is cupric sulfate as a catalyst. The hydrogenation degree can reach over 90%. The hydrogenated product was characterized by IR and DSC. The hydrogenated products behave as a thermoplastic elastomer with excellent thermooxidation resistance, due to the absence of most double bonds and the presence of crystalline domains of polyethylene segments formed by the hydrogenation of polybutadiene segments. Ionomers were obtained by neutralization of the hydrogenated product with metallic ions and characterized by IR, DSC, and TEM. The ionomers also behave as thermoplastic elastomers with mechanical properties better than those of the hydrogenated product without neutralization, due to the existence of ionic domains besides the crystalline domains. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 83: 1375–1384, 2002

Influence of sucrose on cold gelation of heat-denatured whey protein isolate

A solution of heat-denatured whey proteins was prepared by heating 100 g kg−1 whey protein isolate (WPI) at pH 7.0 to 75 °C for 15 min in the absence of salt. Heat treatment caused the globular protein molecules to unfold, but electrostatic repulsion opposed strong protein–protein aggregation and so prevented gel formation. When the heat-denatured whey protein solution was cooled to room temperature and mixed with 15 mM CaCl2, it formed a gel. We investigated the influence of the presence of sucrose in the protein solutions prior to CaCl2 addition on the gelation rate. At relatively low concentrations (0–100 g kg−1), sucrose decreased the gelation rate, presumably because sucrose increased the aqueous phase viscosity. At higher concentrations (100–300 g kg−1), sucrose decreased the gelation rate, probably because sugar competes for the water of hydration and therefore increases the attraction between proteins. These data have important implications for the application of cold-setting WPI ingredients in sweetened food products such as desserts. © 2000 Society of Chemical Industry

Intermediate filament interactions can be altered by HSP27 and alphaB-crystallin.

HSP27 and alphaB-crystallin are both members of the small heat shock protein family. alphaB-crystalllin has been proposed to modulate intermediate filaments and recently a mutation in alphaB-crystallin has been identified as the genetic basis of desmin related myopathy. This disease is characterised in its pathology by aggregates of intermediate filaments associated with alphaB-crystallin. Here we report that HSP27 like alphaB-crystallin is associated with glial fibrillary acidic protein and vimentin intermediate filament networks in unstressed U373MG astrocytoma cells. HSP27 is also associated with keratin filaments in MCF7 cells, indicating that this association is not restricted to a particular intermediate filament type. The association of sHSPs with both the soluble and filamentous intermediate filament fractions of U373 cells was demonstrated biochemically. Heat shock or drug treatments induced a co-collapse of intermediate filaments and associated small heat shock proteins. These data show that the presence of HSP27 or alphaB-crystallin could not prevent filament collapse and suggest that the purpose of this association is more than just filament binding. Indeed, in U373MG cells the intermediate filament association with small heat shock proteins is similar to that observed for another protein chaperone, HSC70. In order to discern the effect of different chaperone classes on intermediate filament network formation and maintenance, several in vitro assays were assessed. Of these, falling ball viscometry revealed a specific activity of small heat shock proteins compared to HSC70 that was apparently inactive in this assay. Intermediate filaments form a gel in the absence of small heat shock proteins. In contrast, inclusion of alphaB-crystallin or HSP27 prevented gel formation but not filament assembly. The transient transfection of GFAP into MCF7 cells was used to show that the induction of a completely separate network of intermediate filaments resulted in the specific association of the endogenous HSP27 with these new GFAP filaments. These data lead us to propose that one of the major functions of the association of small heat shock proteins with intermediate filaments is to help manage the interactions that occur between filaments in their cellular networks. This is achieved by protecting filaments against those non-covalent interactions that result when they come into very close proximity as seen from the viscosity experiments and which have the potential to induce intermediate filament aggregation as seen in some disease pathologies.

Physical properties of cold‐setting gels formed from heat‐denatured whey protein isolate

AbstractCold‐setting gels were formed from a 100 g kg−1 whey protein isolate (WPI) solution (pH 7.0) which had been heated to 90°C for 30 min in the absence of salt. Heating caused the globular protein molecules to unfold and form linear aggregates, but electrostatic repulsion opposed extensive protein‐protein interactions and so prevented gel formation. When the solution containing heat‐denatured proteins was cooled to room temperature and mixed with NaCl (0.2 mol kg−1), the electrostatic charges were shielded and a gel formed. The rate of gel formation increased on cooling, which suggests that the major driving force for protein aggregation and gelation is the hydrophobic effect. Once formed, the rigidity of the gels decreased with decreasing temperature, which suggested that non‐hydrophobic forces (eg configurational entropy, hydrogen bonding, van der Waals forces and disulphide bond formation) are more important in determining the final gel rigidity. The turbidity and rigidity of gels formed by adding salt to preheated WPI at room temperature were less than those of gels formed by heating WPI in the presence of salt. The ability of WPI to form cold‐setting gels with relatively low turbidity extends the range of applications of whey proteins as functional ingredients in the food industry.

Understanding of fundamentals: Key to process modification for tailings reduction

Abstract The tailings produced during bitumen separation from oil sands have a high water holding capacity attributed to ultrafine (≤ 0.2 μm), aluminosilicate clay fractions. These components readily form gels within which both fine and coarse particles may be embedded. This complex mixture (or “fine tails") shows poor dewatering and consolidation characteristics. In this work it has been demonstrated that for gel formation to occur an appropriate combination of ultrafines (amount and particle size) and water chemistry is needed. The natural salt concentration in water recycled from the tailings or pore (connate) water is sufficient to cause the ultrafines to form gels in a matter of days. Gel formers are an integral part of oil sands ores which are unavoidably released during processing. However, one way to prevent gel formation is to change the water chemistry in the extraction process where the bitumen is released. This can be achieved by the addition of sodium silicate instead of sodium hydroxide as process aid in the initial oil sands conditioning step.

Attenuation of immunologic reactivity of lipoprotein(a) by thiols and cysteine-containing compounds. Structural implications.

Samples of human plasma having lipoprotein(a) (Lp[a]) protein levels between 5 and 15 mg/dl and a single apolipoprotein(a) (apo[a]) isoform were incubated in vitro at pH 7.7 with various concentrations (1-20 mM) of N-acetylcysteine, homocysteine, 2-mercaptoethanol (2ME), and dithiothreitol (DTT) for 1 hour at 37 degrees C under a nitrogen atmosphere. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by immunoblot analyses using a polyclonal antibody specific for apo(a) showed a progressive decrease in apo(a) immunoreactivity as a function of reductant concentration. This decrease of apo(a) immunoreactivity was corroborated by enzyme-linked immunosorbent assay (ELISA) using anti-apo(a) as the capture antibody and either anti-apo B or anti-apo(a) as the developing antibody. In turn, there was no significant decrease in the immunoreactivity of apo B-100, as assessed by ELISA using anti-apo B as both the capture and the detecting antibody. In the case of high concentrations of DTT the plasma samples had to be diluted to prevent gel formation on addition of the reductant. A progressive drop in immunoreactivity as a function of reagent concentration was also observed in pure preparations of Lp(a) incubated with the reducing agents at pH 7.7. At equivalent stoichiometries the changes were more marked than those observed with whole plasma, suggesting a quenching effect by the plasma proteins on the activity of the reductants. The changes in immunoreactivity were attended by dissociation of apo(a) from Lp(a) as assessed by Western blotting. This dissociation, which we interpret as the result of cleavage of the interchain disulfide bond(s), was complete at 5 mM DTT and 100 mM 2ME.(ABSTRACT TRUNCATED AT 250 WORDS)