Ion-exchange Fractions Research Articles

CD3 Target Affinity Chromatography Mass Spectrometry as a New Tool for Function–Structure Characterization of T-Cell Engaging Bispecific Antibody Proteoforms and Product-Related Variants

T-cell engaging bispecific antibodies (TCBs) targeting CD3 and tumor-specific antigens are very promising therapeutic modalities. Since CD3 binding is crucial for the potency of TCBs, understanding the functional impact of CD3 antigen-binding fragment modifications is of utmost importance for defining critical quality attributes (CQA). The current CQA assessment strategy requires the integration of structure-based physicochemical separation and functional cell-based potency assays. However, this strategy is tedious, and coexisting proteoforms with potentially different functionalities may not be individually assessed. This increases the degree of ambiguities for defining meaningful CQAs, particularly for complex bispecific antibody formats such as TCBs. Here, we report for the first time a proof-of-concept study to separate and identify critically modified proteoforms of TCBs using functional CD3 target affinity chromatography (AC) coupled with online mass spectrometry (MS). Our method enabled functional distinction of relevant deamidated and glycosylated proteoforms and the simultaneous assessment of product-related variants such as TCB mispairings. For example, CD3 AC-MS allowed us to separate TCB mispairings with increased CD3 binding (i.e., knob-knob homodimers) within the bound fraction. The functional separation of proteoforms was validated using an established workflow for CQA identification based on thoroughly characterized ion-exchange fractions of a 2+1 TCB. In addition, the new method facilitated the criticality assessment of post-translational modifications in stress studies and structural variants in early stage clone selection. CD3 AC-MS has high impact for streamlining the integration of functional and structural characterizations of the large landscape of therapeutic CD3 targeting TCBs from early stage research to late stage characterization.

An arabinan isolated from the antioxidant active fraction of wild blackthorn fruits (Prunus spinosa L.)

Despite the many health benefits of wild blackthorn fruits, their use is negligible and little explored. Preliminary results suggest that this fruit is a rich source of natural antioxidants. Antioxidant active fraction, extracted from wild blackthorn fruits by cold water extraction (Cw), was subjected to ion-exchange chromatography, which gave ten fractions after a successive elution with water, sodium chloride and sodium hydroxide solutions. Fraction eluted by water contained an arabinan of a low molecular weight (Mw = 5000). Its chemical and spectroscopic studies have revealed a highly branched structure with backbone composed of 1,5-linked α-l-arabinofuranose residues; about 76% of them are branched dominantly through O3 and much less through O2, however, some arabinose units (∼8%) were fully branched. In addition to terminal α-L-Araf, the presence of β-L-Araf unit was also identified in blackthorn L-arabinan. Antioxidant activity tests of Cw ion exchange fractions revealed significant effects only of fractions eluted with 1 M sodium chloride (6F) and sodium hydroxide (7F) solutions, while phenolic free arabinan did not possess this effect. Phenolic compounds linked via α-l-arabinosyl side chains appear to be responsible for the antioxidant effect of two Cw fractions.

Leucine-Rich, Potent Anti-Bacterial Protein against Vibrio cholerae, Staphylococcus aureus from Solanum trilobatum Leaves.

A 24 kDa leucine-rich protein from ion exchange fractions of Solanum trilobatum, which has anti-bacterial activity against both the Gram-negative Vibrio cholerae and Gram-positive Staphylococcus aureus bacteria has been purified. In this study, mass spectrometry analysis identified the leucine richness and found a luminal binding protein (LBP). Circular dichroism suggests that the protein was predominantly composed of α- helical contents of its secondary structure. Scanning electron microscopy visualized the characteristics and morphological and structural changes in LBP-treated bacterium. Further in vitro studies confirmed that mannose-, trehalose- and raffinose-treated LBP completely inhibited the hemagglutination ability towards rat red blood cells. Altogether, these studies suggest that LBP could bind to sugar moieties which are abundantly distributed on bacterial surface which are essential for maintaining the structural integrity of bacteria. Considering that Solanum triolbatum is a well-known medicinal and edible plant, in order to shed light on its ancient usage in this work, an efficient anti-microbial protein was isolated, characterized and its in vitro functional study against human pathogenic bacteria was evaluated.

Migration, Transformation Characteristics, and Ecological Risk Evaluation of Heavy Metal Fractions in Cultivated Soil Profiles in a Typical Carbonate-Covered Area

In China, high heavy metal concentrations in cultivated soil are mainly distributed in carbonate-covered areas. The migration and transformation of heavy metals in such soils are influenced by interactions between natural processes and human activities. This study examined the profiles of nine paddy soils, derived from carbonate rocks in Guangxi. The Cd, As, Zn, Cr, Cu, Hg, Ni, and Pb contents we determined, and soil properties such as pH, Corg content, and fractions of Cd, As, Zn, and Cr were tested. Based on the above data, we assessed the vertical distribution of heavy metal fractions, as well as the ecological risks and factors affecting the migration ability of heavy metals, under the influence of human activities and natural soil formation. The results show that compared with the carbonate rocks in Guangxi, the soil profile of the study area is significantly enriched with all eight heavy metals. Among them, Cd, As, Zn, and Cr exceeded China's agricultural land (paddy field) pollution risk screening values, and Cd and As partially exceed the risk intervention values. All fractions of Cd in the soil profiles are widely distributed. The proportion of water-soluble and ion-exchange Cd fractions with high ecological risk decreased significantly from the surface to greater depths in the soil profile. As, Zn, and Cr in the soil profile were mainly in residual states, and the proportion of water-soluble and ion-exchange fractions did not change considerably with increasing depth. The evaluation results of RAC and RSP show that Cd pollution risk in the study area is relatively high, whereas As, Zn, and Cr are generally pollution-free or risk-free. In naturally developed soil in the study area, the clay mineral content and degree of soil development have significant effects on Cd migration, whereas in the cultivated layer affected by human activities, the soil pH and organic matter content are the main controlling factors. The migration ability of soil As is mainly related to soil organic matter, Fe2O3 content, and soil development degree, but organic matter has an obviously enhanced effect in the tillage layer. The main controlling factor of Zn and Cr migration in soil is pH, and the effects are more intense under the disturbance of human activities.

Assessment of mobility and bioavailability of mercury compounds in sewage sludge and composts

Content of heavy metals, including mercury, determines the method of management and disposal of sewage sludge. Excessive concentration of mercury in composts used as organic fertilizer may lead to accumulation of this element in soil and plant material. Fractionation of mercury in sewage sludge and composts provides a better understanding of the extent of mobility and bioavailability of the different mercury species and helps in more informed decision making on the application of sludge for agricultural purposes. The experimental setup comprises the composing process of the sewage sludge containing 13.1mgkg−1 of the total mercury, performed in static reactors with forced aeration. In order to evaluate the bioavailability of mercury, its fractionation was performed in sewage sludge and composts during the process. An analytical procedure based on four-stage sequential extraction was applied to determine the mercury content in the ion exchange (water soluble and exchangeable Hg), base soluble (Hg bound to humic and fulvic acid), acid soluble (Hg bound to Fe/Mn oxides and carbonates) and oxidizable (Hg bound to organic matter and sulphide) fractions. The results showed that from 50.09% to 64.55% of the total mercury was strongly bound to organo-sulphur and inorganic sulphide; that during composting, increase of concentrations of mercury compounds strongly bound with organic matter and sulphides; and that mercury content in the base soluble and oxidizable fractions was strongly correlated with concentration of dissolved organic carbon in those fractions.

The anti-Candida activity by Ancillary Proteins of an Enterococcus faecium strain.

An antimycotic activity toward seven strains of Candida albicans was demonstrated erstwhile by a wild-type Enterococcus faecium isolated from a penguin rookery of the Antarctic region. In the present study the antimicrobial principle was purified by ion exchange and gel permeation chromatography and further was analyzed by LC-ESI-MS/MS. In the purification steps, the dialyzed concentrate and ion exchange fractions inhibited C. albicans MTCC 3958, 183, and SC 5314. However, the gel filtration purified fractions inhibited MTCC 3958 and 183. The data obtained from the LC-ESI-MS/MS indicate that the antimicrobial activity of the anti-Candida protein produced by E. faecium is facilitated by Sag A/Bb for the binding of the indicator organism's cell membrane. Partial N-terminal sequence revealed 12 N-terminal amino acid residues and its analysis shown that it belongs to the LysM motif. The nucleotide sequence of PCR-amplified product could detect 574 nucleotides of the LysM gene responsible for binding to chitin of the cell wall of Candida sp.

Geochemical fractions and risk assessment of trace elements in soils around Jiaojia gold mine in Shandong Province, China

Soils located adjacent to the Jiaojia gold mine were sampled and analyzed to determine the degree of which they were contaminated by trace elements (Hg, As, Cd, Pb, Cu, and Zn) in Shandong Province, China. All 18 samples exhibited mean Hg, As, Cd, and Pb concentrations in excess of local background values, while the mean concentrations of Cu and Zn were below the background values. In addition, the concentrations of trace elements in gold smelter (GS) soils were higher than in the gold mine (GM) soils. The result from a modified Tessier sequential extraction procedure was that with the exception of Cu in soils near the smelter, the trace elements were predominantly associated with the residual fraction. After residual fraction, most Hg was mainly humic acid and strong organic fraction, while most As was the humic acid. Cd was associated with the water soluble, ion exchange, and carbonate fractions compared with the other trace elements. Furthermore, Cu, Pb, and Zn were more concentrated in the humic acid and Fe/Mn oxide fraction. The fractions of trace elements were affected by soil pH and Ec (Electrical conductivity). The humic acid fraction of Hg as well as the ion exchange fraction of Cd and Zn displayed negative correlations with soil pH. The strong organic fraction of Hg, the Fe/Mn oxide fraction of Cd, and the carbonate fraction of Zn were positively related to the soil Ec. The strong organic fraction and ion exchange fraction of Zn were negatively related to soil Ec. However, the ion exchange and carbonate fractions of As showed significant positive correlations with soil pH. A calculated individual availability factor (A f (i) ) is used; the values of each trace element in the soils are in the following order: Cu > Cd > Pb > Zn > As > Hg. When combined with a risk assessment code, data suggest that Hg, As, Pb, and Zn levels showed low risk for the environment, whereas Cd levels in soils adjacent to the GM and Cu levels in soils adjacent to the GS showed medium risk to the environment, and Cd levels in soils adjacent to the GS exhibited higher environment risk.

A broad-spectrum antimicrobial activity of Bacillus subtilis RLID 12.1.

In the present study, an attempt was made to biochemically characterize the antimicrobial substance from the soil isolate designated as RLID 12.1 and explore its potential applications in biocontrol of drug-resistant pathogens. The antimicrobial potential of the wild-type isolate belonging to the genus Bacillus was determined by the cut-well agar assay. The production of antimicrobial compound was recorded maximum at late exponential growth phase. The ultrafiltered concentrate was insensitive to organic solvents, metal salts, surfactants, and proteolytic and nonproteolytic enzymes. The concentrate was highly heat stable and active over a wide range of pH values. Partial purification, zymogram analysis, and TLC were performed to determine the preliminary biochemical nature. The molecular weight of the antimicrobial peptide was determined to be less than 2.5 kDa in 15% SDS-PAGE and in zymogram analysis against Streptococcus pyogenes. The N-terminal amino acid sequence by Edman degradation was partially determined to be T-P-P-Q-S-X-L-X-X-G, which shows very insignificant identity to other antimicrobial peptides from bacteria. The minimum inhibitory concentrations of dialysed and partially purified ion exchange fractions were determined against some selected gram-positive and gram-negative bacteria and some pathogenic yeasts. The presence of three important antimicrobial peptide biosynthesis genes ituc, fend, and bmyb was determined by PCR.

EVALUATION OF HAEMAGGLUTINATION POTENTIALS OF LECTIN ISOLATED FROM ORDEAL TREE (ERYTHROPHLEUM SUAVEOLENS) STEM BARK

Lectin from Erythrophleum suaveolens stem ba rk was partially purified and evaluated in terms of its haemagglutination activity. Ground, dried stem bark of Erythrophleum suaveolens was extracted by aqueous method and the protein extract was concentrated in 20 - 90 % saturation. The ammonium sulfate pre cipitate was subjected to gel filtration chromatography using Sephadex G - 75 resin a nd fractions were tested for hae mag g lutination activity. The two fractions obtained with the highest hemmaglutinating activity were further subjected separately to ion - excha nge chromatography using Sp Sephadex C - 50 resin. I solated lectins were found to agglutinate chicken er ythrocytes; fractions from gel filtration chromatography showed haemagglutinating activity of 0.1 HU/µL and 0.091 HU/µL, while the ion exchange fractions showed haemagglutinating activity of 0.05 HU/µL and all were stable at the temperature range of 30 - 60 0 C and pH range of 3 - 7. The carbohydrate content of the crude extract, Gel I, Gel II, Ion I and Ion II fractions were 0.04 %, 0.04 %, 0.03 %, 0.02 %, respe ctively. The crude and purified lectins inhibited agglutination in the p resence of mannose and N acetyl - glucoseamine sugars and showed five bands at positions 38kDa, 28k Da, 26kDa, 11kDa and 9kDa on a SDS - PAGE electrophoregram.

Molecular heterogeneity of arabinogalactan-protein from Coffea arabica instant coffee

Arabinogalactan-protein complex (AGP), isolated from freeze-dried instant coffee powder of Coffea arabica beans, was subjected to ion-exchange chromatography. Stepwise elution with water and solutions of sodium chloride with increasing ionic strength afforded eight fractions (F1–F8). Chemical analyses have shown that compositions of individual conjugates varied in carbohydrate and protein contents, molecular mass and slightly in monosaccharide composition. Protein content was increasing by increasing ionic strength of the eluent and it was confirmed also by FT-IR spectra. NMR study has shown that carbohydrate moieties in individual ion exchange fractions differed in the degree of backbone and side chains branching. Performed study has confirmed a molecular heterogeneity of arabinogalactan-protein complex present in a commercial instant coffee.

Speciation of heavy metals in paddy soils from selected areas in Kedah and Penang, Malaysia

This study was carried out with the intention of evaluating heavy metal contamination in cultivated paddy areas. The speciation of heavy metals in paddy soils was determined in order to gain insight regarding their origin and distribution in soils. Five sampling sites were randomly selected from Kedah (Yan, Kota Setar, Kubang Pasu district) and Penang (Bumbung Lima district), where the soils constituted marine alluvial deposits. A site in Langkawi, where organic paddy farming is practised was used as the control. The sequential extraction method was adopted in order to obtain the four heavy metal fractions namely the easily leachable and ion exchange (ELFE), acid reducible (AR), oxidizable organic (OO) and resistant (RR) fractions. This study shows that the soil samples were clayey (82 to 96% of grain size Mn>Cr>Cd>Pb>Zn>Cu. Heavy metals such as Pb, Cu, Cr and Zn predominantly occurred in the insoluble form (RR fraction), with the oxides of Fe and Mn incorporated into the clay minerals. Although, the fertilizers and pesticides studied contained low amounts of heavy metals, the elevated amount of amount of Mn and Cd in the soils (ELFE fraction) could possibly be attributed to the longterm and repeated application of these materials to the cultivated paddy areas.

An anticoagulant effect and chemical characterization of Lythrum salicaria L. glycoconjugates

An ion-exchange chromatography of Lythrum salicaria glycoconjugate afforded twelve fractions (LsF1–F12) by stepwise elution with water, sodium chloride and sodium hydroxide solutions. Carbohydrate, phenolic and protein moieties have been determined in all fractions. Individual glycoconjugates LsF1–F12 varied in carbohydrate, phenolic and protein contents, molecular mass and monosaccharide composition. Rhamnogalacturonans and/or homogalacturonans associated with arabinogalactans, arabinan and hexosan types of polysaccharides were identified in ion-exchange fractions. In vitro anticoagulant activity tests showed complete inhibition of plasma clot formation by LsF2, LsF7 and LsF12 fractions and a slight procoagulant effect of LsF3 and LsF4 ones while other ion-exchange fractions were not active.

Proteomics Strategy for Identifying Candidate Bioactive Proteins in Complex Mixtures: Application to the Platelet Releasate

Proteomic approaches have proven powerful at identifying large numbers of proteins, but there are fewer reports of functional characterization of proteins in biological tissues. Here, we describe an experimental approach that fractionates proteins released from human platelets, linking bioassay activity to identity. We used consecutive orthogonal separation platforms to ensure sensitive detection: (a) ion-exchange of intact proteins, (b) SDS-PAGE separation of ion-exchange fractions and (c) HPLC separation of tryptic digests coupled to electrospray tandem mass spectrometry. Migration of THP-1 monocytes in response to complete or fractionated platelet releasate was assessed and located to just one of the forty-nine ion-exchange fractions. Over 300 proteins were identified in the releasate, with a wide range of annotated biophysical and biochemical properties, in particular platelet activation, adhesion, and wound healing. The presence of PEDF and involucrin, two proteins not previously reported in platelet releasate, was confirmed by western blotting. Proteins identified within the fraction with monocyte promigratory activity and not in other inactive fractions included vimentin, PEDF, and TIMP-1. We conclude that this analytical platform is effective for the characterization of complex bioactive samples.

Scavenging and antioxidant activities of immunomodulating polysaccharides isolated from Salvia officinalis L.

Crude polysaccharides, isolated from the aerial parts of sage ( Salvia officinalis L.) by sequential extraction with water (A), hot ammonium oxalate (B), dimethyl sulfoxide (C), 1 M (D) and 4 M (E) potassium hydroxide solutions, and six ion-exchange fractions of A were examined for their ability to inhibit peroxidation of liposome lipid by hydroxyl radicals and to reduced DPPH radical content. The highest inhibition of liposome lipid peroxidation was found with crude polysaccharides A, B and D, antioxidant activities reached ∼37%. The purified fractions A1 and A2 inhibited the liposome peroxidation to ∼35%. However, the radical scavenging abilities of the most active crude polysaccharides A, B and C on DPPH radicals were found in the range 80–90%, while the most active purified fractions A 3–A 6 in three or fourfold doses achieved 75–92%. The least effective tested polysaccharides succeeded 20% inhibition using both methods.

3557 POSTER Phase I study of docetaxel, oxaliplatin and S-1 (DOS) for patients with advanced gastric cancer

Content of heavy metals, including mercury, determines the method of management and disposal of sewage sludge. Excessive concentration of mercury in composts used as organic fertilizer may lead to accumulation of this element in soil and plant material. Fractionation of mercury in sewage sludge and composts provides a better understanding of the extent of mobility and bioavailability of the different mercury species and helps in more informed decision making on the application of sludge for agricultural purposes. The experimental setup comprises the composing process of the sewage sludge containing 13.1 mg kg−1 of the total mercury, performed in static reactors with forced aeration. In order to evaluate the bioavailability of mercury, its fractionation was performed in sewage sludge and composts during the process. An analytical procedure based on four-stage sequential extraction was applied to determine the mercury content in the ion exchange (water soluble and exchangeable Hg), base soluble (Hg bound to humic and fulvic acid), acid soluble (Hg bound to Fe/Mn oxides and carbonates) and oxidizable (Hg bound to organic matter and sulphide) fractions. The results showed that from 50.09% to 64.55% of the total mercury was strongly bound to organo-sulphur and inorganic sulphide; that during composting, increase of concentrations of mercury compounds strongly bound with organic matter and sulphides; and that mercury content in the base soluble and oxidizable fractions was strongly correlated with concentration of dissolved organic carbon in those fractions.

Effect of diffusion and resuspension on nutrient release from submerged sediments

Nutrient release from a coarse-sand (CS) and a fine-sand (FS) estuarine sediment into the overlying water column was studied under static (diffusion-controlled) and dynamic (resuspension) conditions. Resuspension increased the amounts of and P released from the sediment cores compared to those released by diffusion alone due to direct release of these nutrients from the readily available pore-water and ion exchange fractions. The average effective and P diffusion coefficients (D e) were found to be similar for both sediments, and did not vary significantly between static and dynamic conditions. Values of D e were typically of the order of 10−5 to 10−6 cm2 s−1. The average daily and P diffusive flux density from CS and FS into the overlying water column of the study area was estimated to be 24.6, 0.02 and 0.39 mg m−2 d−1, and 66.9, 0.017 and 0.002 mg m−2 d−1, respectively. These fluxes suggest that nutrient release from the sediments via diffusion and small-scale resuspension events may represent only a minor source of and P to the water column of the study area.

CHARACTERIZATION OF PHYTOCHELATIN-LIKE COMPLEXES FROM FLAX (LINUM USITATISSIMUM) SEED

Proteins were extracted from dehulled and delipidated powder of flaxseed (cv. NorMan), and fractionated by DEAE-Sephacel anion exchange chromatography. Ultraviolet absorbance scans of ion exchange fractions eluting in buffers at pH 8.6 containing 0. 45 M and 0.50M NaCl revealed A 255 >A 280 . Furthermore, the A 255 was reduced upon acidification and restored by adjusting to the original pH, confirming spectral characteristics typical of metal-thiolate complexes. Characteristic circular dichroism spectra and reversible aggregation-dissociation behavior under low and high salt conditions observed for these fractions are also typical of metal binding phytochelatins reported for other plants, while the presence of aromatic amino acids indicated by fluorescence spectra suggests some similarity to nonmetallothionein proteins. Further work is underway to investigate the prevalence of these and other complexes as well as the distribution of metals in different protein fractions of flaxseed, as a function of cultivar and geographical location.

Characterization of a new family of toxin-like peptides from the venom of the scorpion Leiurus quinquestriatus hebraeus. 1H-NMR structure of leiuropeptide II.

To extend our knowledge about the structural features of short scorpion toxins, the ion-exchange fractions obtained from Leiurus quinquestriatus hebraeus venom were investigated by plasma desorption mass spectrometry in order to select low molecular mass polypeptides. Three toxin-like peptides with molecular mass close to 3 kDa, named leiuropeptides I, II and III, were purified and found devoid of any significant toxicity against mammals and insects. Their amino acid sequences revealed a cysteine pattern analogous to that of short-chain scorpion toxins. The solution structure of leiuropeptide II was determined by 2D 1H-NMR spectroscopy and indicated the presence of a helix accommodating a proline, connected to a two-standard beta-sheet by three disulfide bonds. The overall fold of leiuropeptide II is found to be similar to that of leiurotoxin I, a 31-residue toxin present in the same scorpion venom which acts on K+ channels. In order to rationalize the absence of toxicity, the electrostatic potential of leiuropeptide II was compared to that of leiurotoxin I. The peptide is characterized by a large negative zone around Glu4, Asp5 and Asp8 residues, beginning in the neighbourhood of the beta-turn and extending along the helix. In the same area, leiurotoxin I exhibits a positive surface, around Arg6 and Arg13 basic residues, which are essential for its receptor affinity.

Albumin adducts in plasma from workers exposed to toluene diisocyanate.

Desalted plasma from a 2,4- and 2,6-toluene diisocyanate (2,4- and 2,6-TDI) exposed worker at a factory producing flexible polyurethane foam was separated and fractionated into 200 fractions using ion-exchange chromatography followed by a gel-filtration separation and fractionation into 59 fractions. The corresponding amines (to the isocyanates), 2,4- and 2,6-toluenediamine (2,4- and 2,6-TDA), were determined in each fraction after sulfuric acid hydrolysis as pentafluoropropionic anhydride derivatives by capillary gas chromatography and chemical ionisation mass spectrometry monitoring negative ions. The ion exchange fractions containing TDA (81-115) were added together and the solution was separated and fractionated on the gel-filtration column. The fractions 81-115 contained 84 and 72% of 2,4- and 2,6-TDA, respectively, as compared to the unfractionated plasma. The gel filtration fractions 22-27 contained 107 and 119% of 2,4- and 2,6-TDA, respectively, as compared to the amounts in the ion exchange fractions (81-115). Agarose gel-electrophoresis and electroimmunoassay demonstrated that albumin, 2,4- and 2,6-TDA co-eluted in both ion-exchange and gel-filtration chromatography. Quantitative determination of albumin, 2,4- and 2,6-TDA also demonstrated that these components co-eluted using albumin-immunosorption chromatography. In addition, studies of affinity isolated IgG revealed that this fraction was devoid of 2,4- and 2,6-TDA. These results indicate that albumin is the major receptor molecule for 2,4- and 2,6-TDI in blood plasma and that these isocyanates form covalent bondings with albumin.

POLARITIES OF SHRP ASPHALTS CALCULATED FROM ION EXCHANGE CHROMATOGRAPHY SEPARATION DATA

Abstract During the Strategic Highway Research Program, four asphalts were separated into polar and non-polar fractions by means of three different ion exchange chromatography procedures. For each asphalt, the non-polar neutral fractions comprised over 50 mass percent of the total asphalt. These neutral materials are much less viscous than corresponding whole asphalts. Molecular weights of the neutral fractions correlate roughly with viscosities, whereas there is no correlation between these properties for whole asphalts. The various polar fractions were subdivided into acidic, basic, and amphoteric fractions of differing polarities. Based on dipole moments of typical components, polarity factors were assigned to each of these fractions. Multiplication of mass percentages of each fraction by their polarity factors yields polarity indices. Total polarities of asphalts or mixtures of ion exchange fractions are obtained by summing polarity indices of the unique distribution of components. Polarities of aspha...