Department of Chemistry and the MOE Key Laboratory of Spectrochemical Analysis and Instrumentation, College of Chemistry and Chemical Engineering,Xiamen University, Xiamen, 361005, China
Core-shell particle is a new generation high performance packing material for liquid chromatography.Through comparison with a classical totally porous silica phase of the same particle size, we studiedAscentis Express 5 mm core-shell particle's electrochromatographic behavior, in terms of voltage-currentproperty, electroosmotic flow (EOF) and van Deemter curve. It was found, due to the nonpermeable solidcore, the core-shell particle presented a diminished EOF and efficiency than the totally porous paricle.This on the other hand proved that the intra-particle pore flow extensively exists and plays an importantrole in electrochromatography on totally porous material. The core-shell particle's high retentivity led toan enhanced resolution for weakly retained hydrophilic peptides, which were poorly retained and co-eluted on totally porous particles. Further exploration has shown the core-shell material can achieveefficient electrochromatography of protein digests, excellent performance in terms of resolution,reproducibility and long term stability have been observed. The results indicate that the core-shellstructure may suggest a reasonable design of stationary phase for bioelectrochromatography of pep-tides and proteins.
Core-shell particle was recently reintroduced as a high perfor-mance chromatographic material [1e4]. It has excellent masstransfer property as well as high uniformity in size distribution, in
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comparison with conventional totally porous particulate material[5,6]. Core-shell chromatographic material has won extensiveacceptance in many fields, such as pharmaceuticals, environmentalscience and biomedical research [7]. So far, the greatest advantageof core-shell material that has been demonstrated is its high res-olution realized at high speed on HPLC instrumentation [5e7]. Thismade it competent with UHPLC technology which needs sub-2 mmmaterial and ultra-high pressure instrumentation to gain highresolution and high speed.
In the past decade, a series of core-shell material has beenintroduced into the market, such as Poroshell by Agilent, Halo byAMT, Ascentis Express by Sigma-Aldrich and Kinetex by Phenom-enex etc. [8e18]. It is exciting to notice that, apart from the2.6e2.7 mm particles introduced in the early days, recently both5 mm and sub-2 mm core-shell particles have become commerciallyavailable [12,13,19]. Lately, wide-pore core-shell particles withparticle sizes of 3.4e3.6 mm and pore sizes of 160e400Å, targetingat HPLC of biomolecules, have also been released [14,20]. Mean-while, fundamental and applied studies focusing on this newgeneration HPLC material, have been extensively reported [4,5,7].
Apart from HPLC, capillary electrochromatography (CEC) is alsoa high performance liquid chromatographic technique but drivenby electroosmotic flow (EOF) [21e23]. It holds unique selectivity forcharged molecules due to its chromatography-electrophoresiscombined separation mechanism. Over the years, however, CEC isstill struggling to become a robust and routine separation tool.Although different viewpoints have been discussed, one pointlargely agreed is that it was the lack of proper stationary phasesspecifically designed for CEC that holds the advancement of elec-trochromatography [24].
In this paper, we studied electrochromatographic behavior ofthis new generation core-shell material to find out: its feasibilityand suitability as a CEC medium; its electrochromatographic fea-tures in terms of EOF, separation efficiency as well as retentionproperties; finally and most importantly whether this new type ofmaterial is of usefulness in electrochromatography of peptides andprotein digests, in order to explore its effectiveness in microscalebioseparations.
2. Experimental
2.1. Materials and apparatus
Fused silica capillaries, 100 mm i.d., 365 mm o.d. and 20 mm i.d.,90 mm o.d., were purchased from Yongnian Reafine Chromatog-raphy (Hebei, China). Perfusive silica beads, 110 mm in diameter,pore size ~1 mm, to be used as frits of capillary columns, were ob-tained from X-tec (Bromborough, UK). Core-shell particulate ma-terial was from an Ascentis Express C18 column, 5 mm, 90Å,purchased from Sigma-Aldrich (St. Louis, MO). Totally porousreversed phase packing material, Waters Spherisorb ODS, 5 mm,300Å, from Waters (Milford, MA) were used for capillary columnpacking. Tris-(hydroxymethyl)aminomethane hydrochloride (Tris-HCl, biochemical grade), NH4HCO3, thiourea, methyl-, ethyl-, pro-pyl-, and butylbenzenes of analytical grade, dithiothreitol (DTT),iodoacetamide (IAA), trifluoroacetic acid (TFA), trypsin ofsequencing grade and a standard protein cytochrome C were pur-chased from Sigma-Aldrich (St. Louis, MO). Acetonitrile (ACN) andacetone of HPLC grade were provided by Merck (Darmstadt, Ger-many). Ultrapure water (18.2MU) was prepared in a Milli-Q system(Millipore Filter Co., Bedford, MA) and used throughout this study.All chemicals and reagents were at least of analytical or high gradeand used without further purification. An Elite P230 high pressurepump from Dalian Elite Analytical Instruments (Dalian, China) wasused for column packing. A Waters Quanta 4000E capillary
electrophoresis system (Waters, Millford, MA) equipped with asingle wavelength UV (214 nm) detector, without thermostattingand pressurization facilities, was used for electrochromatographyseparations.
2.2. Protein digestion
A complex peptide mixture was prepared by tryptic digestion ofcytochrome C in solution. The protein sample was solubilized in8M urea, 50mM NH4HCO3. Then, the sample was reduced by DTTand alkylated by IAA. Finally, trypsin was added at a protein-to-enzyme ratio of 50:1, the digestion was incubated at 37 �C overnight. The protein digest sample was desalted and concentratedusing a SPE C18 column (Thermo Scientific, Bellefonte, PA).
2.3. Column preparation
The capillary columns used in this study were prepared ac-cording to a single particle fritting technology as we previouslyreported [25e30]. In brief, a detectionwindow for optical detectionwas first created by burning off a short segment of polyimidecoating of an empty fused silica capillary, using a resistance wire-based capillary burner. Then a single perfusive particle, ~110 mmin diameter, was tapped in from the short end of the capillary. Anarrow bore capillary, 90 mm o.d. was used as a plunger to push theparticle inward to a position right past the detection window. Thissingle particle was used as the outlet end frit of the column. Then,from the other end of the capillary, a slurry of the packing materialwas introduced and packed under a high pressure up to 6000 psi.When the column bed was packed to the length, another singleparticle was tapped in as the inlet end frit.
Capillary columns of 100 mm i.d., 365 mm o.d., with a packedsection of 20 cm and total length of 28 cm, packed with AscentisExpress C18 5 mm core-shell particles, and Waters Spherisorb C185 mm totally porous particles, respectively, were fabricated based onthemethod described above. Following fabrication, the columnwashydraulically equilibrated with the mobile phase using a manualsyringe pump for a certainwhile andmounted onto the instrument.The column was then electrically equilibrated with the same mo-bile phase under applied voltages of 5, 10, 15, 20, 25 and 30 kVsequentially. After equilibration, the column was ready for use.
2.4. Electrochromatographic separations
For CEC separation of aromatic hydrocarbons, the mobile phasewas ACN/aqueous Tris-HCl (50mM, pH 8.5), 80:20 v/v. The sampleused for van Deemter curve evaluation was a mixture of alkyl-benzenes (methyl-, ethyl-, propyl-, butyl- and amylbenzenes) withthiourea (as the dead time marker) dissolved in the mobile phase,with each component having a concentration of ~0.5mM. For CECseparation of polycyclic aromatic hydrocarbons (PAH), the samplewas a mixture of 16 PAHs, primary pollutants designated by theEPA: naphthalene, acenaphthylene, acenaphthene, fluorene,phenanthrene, anthracene, fluoranthene, pyrene, benz[a]anthra-cene, chrysene, benzo[b]fluoranthene, benzo[k]fluoranthene,benzo[a]pyrene, dibenz[a,h]anthracene, benzo[ghi]perylene, andindeno[1,2,3-cd]pyrene. Another PAH mixture of naphthalene,acenaphthene, anthracene and pyrene was also prepared and dis-solved in the mobile phase, with each component having a con-centration of ~0.5mM. The samples were introducedelectrokinetically at 10 kV for 5 s.
For CEC separation of standard peptides and protein digests, themobile phase was ACN/aqueous phosphate (50mM, pH 2.5), 40:60v/v. The peptide sample solution was a mixture of Gly-Tyr, Val-Tyr-Val, Met enkephalin, Leu enkephalin and angiotensin II dissolved in
Fig. 1. Electrochromatography of alkylbenzenes on six capillary columns packed withcore-shell particles. Experimental conditions: CEC columns, 200mm� 100 mm i.d.(total length 280mm); packed with Ascentis Express 5 mm C18; mobile phase, ACN/Tris-HCl (50mM, pH 8.5), 80:20 v/v; applied voltage, 10 kV; analytes: thiourea,methyl-, ethyl-, propyl-, butyl- and amylbenzenes (in order of elution); electrokineticinjection: 10 kV/5 s; UV detection: 214 nm.
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the mobile phase, with each component having a concentration of~0.1mgmL�1. All the samples were introduced electrokinetically at10 kV for 10 s. The CEC separations were performed at a roomtemperature of ~25 �C.
3. Results and discussion
3.1. Fabrication of columns packed with core-shell particles
The conventionally used fritting method for capillary columnfabrication is based on sintering [31e33]. Both the sintered frit andthe packing material are totally porous silica, who have the similarchemistry and porosity. While the core-shell material has adifferent structure, it has a solid core and a superficially porousouter layer. Such a core-shell particle-packed bed has been foundhard to be fritted through sintering the core-shell particle itself[34]. Therefore, initial attempts on making core-shell capillarycolumns have turned to sintering porous silica as frits or usingmonolithic frits [35e37]. This of course introduced nonuniformityto the packed bed, especially not ideal for electrochromatography,whose driving force (i.e. EOF) is sensitive to the packed bed surface.
We introduced a single particle fritting technology for capillarycolumn fabrication [25e30]. Since it is based on a purely physicalprocess, keystone effect, it does not need sintering to frit. Therefore,the single particle fritting technology has an excellent applicabilityto different types of packing material. In this study, the core-shellparticle packed bed was fritted at both ends using single particlefrits. The frit has a very short length of ~100 mm, greatly diminishedthe nonuniformity of the packed bed.While sintered frits inevitablyhave a long frit length, usually 2e5mm [31,32], whichmay give riseto dead volume and nonuniformity to the core-shell particle-packed bed.
To investigate the repeatability of column preparation based onthis single particle fritting technology, we fabricated six capillarycolumns packed with core-shell particles. Using an alkylbenzenemixture as the probe, we evaluated their column-to-columnreproducibility. As shown in Fig. 1, electrochromatograms withgood consistency were recorded. Using amylbenzene (the lastpeak) as the standard, a relative standard deviation of retentionfactor at 0.6% was obtained, indicating an excellent column-to-column reproducibility.
3.2. Electrochromatographic performance: core-shell vs. totallyporous particles
The electrochromatographic performance of core-shell particleswas evaluated through comparison with a totally porous silicaphase. It needs to be highlighted that in this study, in order to havea head-to-head comparison, we intentionally chose a 5 mmAscentisExpress core-shell particle material to compare with a 5 mm totallyporous particle material, although other size core-shell particles,e.g. 1.7, 2.6, 2.7 and 3.6 mm have been extensively reported [8e18].The Ascentis Express core-shell C18 particle has a nominal particlesize of 5 mm, pore size of 90Å. A totally porous silica phase, WatersSpherisorb C18, 5 mm, pore size 300Å, was chosen for comparison.
We first investigated the electric properties of the two phases.As shown Fig. 2A, the two phases have discrepant voltage-currentcurves. Under the same applied voltage, the core-shell particle-packed bed presented a lower conductivity, in this case, around 2times lower than the totally porous particle-packed bed. This ismainly because the core-shell particle has a large solid core, wherethe electrolyte cannot diffuse in and contribute to the total con-duction. As evidenced in the TEM graph (Fig. 3), the nominal 5 mmcore-shell particle has an actual particle size of around 5.9 mm, anda thin porous outer layer of 0.2e0.5 mm, i.e. the spherical particle is
largely solid. The electrolyte can only occupy the inter-particlespace of the packed bed and the thin porous layer of the core-shell particles. While for the totally porous material, the electro-lyte can diffuse into the intra-particle pores, apart from the inter-particle space. According to Giddings [38], a randomly packedbed has an inter-particle porosity of around 0.4. In other words, anon-porous particle-packed bed has a total porosity of ~0.4.While atotally porous particle-packed bed normally has a total porosity of~0.8, including inter-particle and intra-particle pores [39,40]. If wetreat the core-shell particle as a solid sphere and take 0.4 as its totalporosity, in comparison with the totally porous particle, thenaround two times difference in electric resistance is a reasonableresult. This fits well with the experimental data shown in Fig. 2A.This indicates the conductive contribution from the superficialpores of the core-shell particle is not significant. In comparisonwith the 5 mm totally porous particle, this thin porous layer did notelectrically contribute much to the bed's permeability.
We further evaluated electroosmotic flows of the core-shell andtotally porous particle-packed columns. As shown in Fig. 2B, thetotally porous particle presented higher EOF than the core-shellparticle. Due to its unpermeable core, the core-shell particle-packed bed's EOF should solely come from the inter-particle space.While for the totally porous phase, there are rich pores throughoutthe porous particle. According to Rice andWhitehead [41], the ratioof pore size to electric double layer thickness determines theprobability of intra-particle (pore) flow. When this ratio is over 20,then 80% EOF is allowed, i.e. there will be significant intra-particleEOF [42,43].
Fig. 2. Performance comparison of core-shell and totally porous particle packed columns. A: voltage-current curves; B: EOF as a function of applied voltage; C: EOF as a function ofelectrolyte pH; D: van Deemter curves. Both columns are 200mm� 100 mm i.d. (total length 280mm); packed with Ascentis Express 5 mm C18, and Waters Spherisorb 5 mm C18particles, respectively, as indicated in figure; mobile phase in A, B and D: ACN/Tris-HCl (50mM, pH 8.5), 80:20 (v/v); mobile phases used in C: NaH2PO4-H3PO4 (pH 3.0, 4.0, 5.0),Na2HPO4-NaH2PO4 (pH 6.0, 7.0), Na2B4O7-H3BO3 (pH 8.0, 9.0), 25mM, containing 40% (v/v) ACN, respectively. In B, C and D: electrokinetic injection, 10 kV/5 s; UV detection,214 nm; Analytes: DMSO, as the dead time marker for EOF; amylbenzene, as the probe for peak efficiency.
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d¼(ε0εrRT/2 cF2)1/2 (1)
Quantitatively, equation (1) was used to estimate the doublelayer thickness, d, where ε0 is the permittivity of a vacuum, εr is thedielectric constant, R is the universal gas constant, T is the absolutetemperature, c is the electrolyte concentration, F is the Faradayconstant. In this case, the mobile phase is ACN/Tris-HCl (50mM, pH8.5), 80:20 (v/v). The overall electrolyte concentrationc¼ 0.010mol L�1, and ε0¼ 8.85� 10�12 C2 N�1m�2, εr¼ 39.26,R¼ 8.3145 J K-1 mol-1, T¼ 298 K and F¼ 96500 Cmol�1. The dielec-tric constant value was taken from reference [44,45]. The doublelayer thickness was calculated to be 21.5Å. Although the 5 mmtotally porous particle's nominal pore size is 300Å, the experi-mentally determined pore size distribution is quite wide, as shownin Fig. 4, it spanned from 50 to >700Å. Therefore, in practice, theratio of pore size to double layer thickness may reach 32. In com-parisonwith the 5 mm core-shell particles, it is clear that the totallyporousmaterial, under this electrolyte condition, can support intra-particle EOF, which contributed significantly to the overall EOF, asevidenced in Fig. 2B.
We further extended the evaluation of EOF. As shown in Fig. 2C,from pH 3e9, the totally porous particle-packed column has pre-sented higher EOF than the core-shell particle-packed column.Based on the electrolytes used in Fig. 2C, the overall electrolyteconcentration is 15mM and the ACN concentration is 40%. There-fore, in this case, c¼ 0.015mol L�1, ε0¼ 8.85� 10�12 C2 N�1m�2,εr¼ 50.77, R¼ 8.3145 J K-1mol-1, T¼ 298 K and F¼ 96500 Cmol�1,the double layer thickness was calculated to be 20.0Å.
According to the data shown Fig. 4, the totally porous materialhas a considerable portion of wide pores: about 23% of the totalpore volume is of the pores greater than 400Å. In other words, itwell satisfied the ratio of 20 times, supporting significant intra-particle EOF on the totally porous particle-packed column. Whilefor the core-shell material, its unique solid core structure ruled outthe possibility of intra-particle EOF. In this regard, the core-shellparticle can be used as a standard material which has zero intra-particle pores and therefore, zero pore-flow. It needs to be notedthough, as Fanali et al. pointed out [46], theremay still be tangentialintra-particle EOF at the superficial porous layer of core-shell par-ticles. In our case, however, as visualized in Fig. 3, the superficial
Fig. 3. TEM graph of the 5 mm core-shell particles. The scale bar is 2 mm.
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porous layer is merely ~0.2e0.5 mm thick. What's more, the su-perficial pore size, according to the manufacturer [19], is 90Å,which is too narrow to support the pore-flow. Nevertheless, wenoticed that, recently AMT introduced Halo Protein & Peptide core-shell media [20], which have nominal pore sizes of 400 and 160Å,respectively. Such wide pores in theory may support significantsuperficial pore-flow on the particle surface.
We further investigated van Deemter curves of the two phases(Fig. 2D), as in theory improved chromatographic dynamics should
Fig. 4. BET characterization of the totally p
be expected due to the pore flow-enhanced mass transfer. Usingthe core-shell particle-packed column as the benchmark, our dataindicates, the totally porous particle-packed column has betterperformed dynamics, especially in the high speed range, as shownin Fig. 2D. This is a clear sign of the pore flow-enhanced masstransfer and therefore lower plate heights. Meanwhile, it is worthnoting that both curves have a relatively flat pattern at the highflow rates, i.e. both particles support a good mass transfer. For thetotally porous particle, this should be due to the intra-particle EOF,which support a convective instead of diffusive mass transfer.While for the core-shell particle, this should be attributed to thesimplified mass transfer path, i.e. the analytes do not need toexperience a deep diffusion path, due to the superficial pores [4,7].
3.3. Electrochromatography of neutral compounds, peptides andprotein digests
The core-shell column was first applied to electro-chromatography of polycyclic aromatic hydrocarbons (PAHs). Asshown in Fig. 5, the core-shell stationary phase presented strongeroverall retention of the PAHs than the totally porous phase. Wethen chose six PAHs for further analysis, as shown in Fig. 6.Quantitatively, the retention factors of the PAHs on the core-shellphase are around 3 times of that on the totally porous phase. Thetotally porous phase used here has a carbon content of 10.8% [47],while no data on the carbon content of the Ascentis Express core-shell particles was provided. In the published data, a carbon con-tent of 12% was reported for Kinetex C18 particles, another similarcore-shell particulate chromatographic material from Phenomenex[18,48]. This indicates that technologically, core-shell material'ssuperficial pores can be effectively functionalized with C18 groupsand providing hydrophobicity. However, as shown in Figs. 5 and 6,in comparisonwith the totally porous C18 phase, the core-shell C18phase presented a significantly higher hydrophobic retention. This
Fig. 6. Retention factors of six PAHs separated on the core-shell and totally porousparticle-packed columns. The six PAHs are: (1) acenaphthylene, (2) pyrene, (3) amyl-benzene, (4) chrysene, (5) benzo[a]pyrene, (6) dibenz[a,h]anthracene. Experimentalconditions as in Fig. 5.
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may be due to the pore structure and more importantly, theiraccessibility during the chromatographic process. McDonald had anexcellent discussion about particulate reversed-phase material andpore grading according to their size, depth and accessibility [49]. Itwas suggested that, the wide (>100Å) and near surface pores arewell C18-functionalized, while the smaller and deeper pores cannotbe effectively functionalized with long-chain silyl bonding reagents(e.g. C18 groups), although they may still be accessed by analytes.The core-shell material's pores locate at the superficial layer, in thiscase, around 0.2e0.5 mm deep, therefore the C18 group can be wellloaded in during the functionalization process. While for the totallyporous material, the pores can distribute throughout the particleand generally have a greater depth. The smaller (e.g. <100Å, assuggested by McDonald [49]) and/or deeper pores cannot be aseffectively C18-bonded as the wide and near surface pores duringthe functionalization process, resulting in an uneven distribution ofC18 groups throughout the spherical particles. Overall, the analyte-accessible surface of core-shell particle is relatively higher carbon-loaded than that of the totally porous particle. This retentivity
difference is especially significant in the electrochromatographymode. As in HPLC mode, the totally porous material has no intra-particle flow, those (deep) diffusion pores are not accessible.While in CEC mode, the electroosmotic intra-particle flow existsand enables chromatographic interactions to take place in thosedeep diffusion pores, whose carbon load is significantly lower thanthe average level. Consequently, the apparent retention on thetotally porous phase is weakened.
When applied to CEC of peptides, the core-shell column pre-sented an excellent retentivity and resolution, as shown in Fig. 7.The oligopeptides, Gly-Tyr, Val-Tyr-Val and Met enkephalin, werenot effectively retained and co-eluted at 6min on the totally porouscolumn. Whilst the same oligopeptides, were well retained andresolved on the core-shell column in a time window from 8 to12min. Both peak width and peak capacity are better on the core-shell column than on the totally porous column. Althoughdemonstrated with standard peptides, it needs to be highlightedthat, such weakly retained peptides tend to co-elute around thedead time on C18 columns, causing information loss in proteinidentification in proteomics analyses. In this regard, the core-shellphase may find a good use in retaining and effectively resolvinghydrophilic peptides in reversed phase chromatography. On theother hand, the core-shell material may be an ideal solid phaseextraction medium used as trap columns in front of LC-MS, in orderto trap peptide species at a high recovery, while filtering out saltsand other small molecules in omics research.
Further, we applied the core-shell material to electro-chromatography of a protein digest cytochrome C. As shown inFig. 8, again, the core-shell column achieved a stronger retentionand better resolution than the totally porous column, suggesting itsusefulness in electroseparation of complex mixtures of peptides. Itneeds to be pointed out, however, the separation conditions usedfor electrochromatography of tryptic digest of cytochrome C werenot optimized, when gradient elution conditions [50,51] areapplicable, the resolution may still be improved.
The column was used for one week to investigate its long termstability and reproducibility. Reproducible electrochromatogramsof the protein digest have been recorded, as shown in Fig. 9. Whilein the same time window, degrading electrochromatograms havebeen observed on the totally porous particle-packed column (datanot shown). This should be due to the rich intra-particle surface
Fig. 7. Electrochromatography of standard peptides on core-shell (A) and totallyporous (B) particle-packed columns. Experimental conditions: CEC columns,200mm� 100 mm i.d. (total length 280mm); packed with Ascentis Express 5 mm C18,and Waters Spherisorb 5 mm C18 particles, respectively; mobile phase, ACN/phosphate(50mM, pH 2.5), 40:60 v/v; applied voltage, 10 kV; sample: Gly-Tyr, Val-Tyr-Val, Metenkephalin, Leu enkephalin and angiotensin II (in order of elution); electrokineticinjection: 10 kV/10 s; UV detection: 214 nm.
Fig. 8. Electrochromatography of tryptic digest of cytochrome C. Experimental con-ditions as in Fig. 7.
Fig. 9. Electrochromatography of tryptic digest of cytochrome C. As specified, the threeelectrochromatograms were recorded in a time span of one week. Experimentalconditions: CEC columns, 200mm� 100 mm i.d. (total length 280mm); packed withAscentis Express 5 mm C18; mobile phase, ACN/phosphate (50mM, pH 2.5), 40:60 v/v;applied voltage, 10 kV; electrokinetic injection: 10 kV/10 s; UV detection: 214 nm.
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area: on one hand it enhanced the EOF, speeding up CEC separa-tions; on the other hand, however, the surface tended to becontaminated through adsorption of biomolecules, in this case,tryptic peptides. Consequently, the surface lost its activation forEOF and resulted in irreproducible CEC and degrading separationperformance. While the core-shell particle, due to its large solidcore and limited pore surface, greatly diminished the probability ofbiocontamination, maintaining a relatively stable charged surfacefor EOF and good performance of bioelectrochromatography. Thecore-shell structured material may suggest an ideal stationaryphase design for (bio)electrochromatography and has a good po-tential in CEC-MS for proteomics.
4. Conclusions
Core-shell particles were evaluated for their performance inelectrochromatography. The solid core plays a key role in theelectric properties of core-shell particles. Using the core-shell ma-terial as a benchmark, it is clear that the totally porous material'sperformance in electrochromatography is supported by a signifi-cant intra-particle pore flow. The pore flow provided a higher EOFand improved mass transfer, therefore enhanced the separationefficiency. In contrast, the core-shell particle's unpermeable solidcore completely eliminated the pore flow. In terms of chromato-graphic performance, the core-shell material presented a strongreversed-phase retention and improved resolution especially forweakly retained (hydrophilic) species. Electrochromatography ofstandard peptides and protein digests has shown excellent sepa-rations of hydrophilic peptides and an improved overall resolution.The non-porous core minimized the risk of biocontamination ofintra-particle pores, who are also responsible for EOF generationand may lead to performance degradation upon biocontamination.The solid core structure rules out such uncertainty and maintaineda long term stability of electrochromatography. These unique ad-vantages of the core-shell material may find applicabilities in bio-electrochromatography and CEC-MS for proteomics.
Acknowledgements
This work was supported by National Natural Science
L. Yang et al. / Analytica Chimica Acta 1033 (2018) 205e212212
Foundation of China (21475110, 21535007), Fundamental ResearchFunds for Central Universities of China (20720150161,20720160051), Xiamen Science and Technology Project(3502Z20173019), NFFTBS (J1310024) and PCSIRT (IRT13036).
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