Compressed fluids in analytical separation methods Michal Roth Department of Fluid Phase Separations Czech Academy of Sciences, Institute of Analytical Chemistry, v. v. Veveří 97 60200 Brno employing the results and presentations by Pavel Karásek, Josef Planeta, Elena Varaďová Ostrá, Jaroslav Pól, Barbora Hohnová, Lenka Šťavíková, Marie Horká, Dana Moravcová and Karel Šlais Structure topic outline - why compressed fluids in separations ? 1) supercritical fluid chromatography (SFC) 2) supercritical fluid extraction (SFE) 3) extraction with organic solvents at elevated Tand P PFE - Pressurized Fluid Extraction - Pressurized Liquid Extraction PLE-PSE-ASE- Pressurized Solvent Extraction Accelerated Solvent Extraction 4) extraction with pressurized hot (subcritical) water PHWE - Pressurized Hot Water Extraction SubWE - Subcritical Water Extraction 5) supercritical water vs. siliceous surfaces - application in analytical separations SFE, SFC £p SOLID LIQUID °"""Si"Fr- SB t SF - supercritical fluid ' GAS —- teplota t — utilization of fluid properties above their respective Tc and Pc — properties (density, solvent power,...) controlled by P and T — COz Tc ~ 31 °C Pc~7.8 MPa — density, solvent power ~ liquids — viscosity, diffusion rate ~ gases —faster than liquid chromatography/extraction — environment-saving — less organic solvents (or none at all) 1) Supercritical fluid chromatography (SFC) * SFC apparatus * Preparation of columns for SFC (micro HPLC) * Examples of SFC separations * Non-analytical applications - systems with ionic liquids Příprava kapilárních náplňových kolon pro SFC (HPLC) Požadavky na kolony: Náplň sorbent o zrnitosti 3 nebo 5 um, délka kolony do 1m • Průměr kolony do 320 um => F = 4[il/min(liq.), F = 10ml/min(g) • Pracovní tlak do 40 MPa => nároky na uzavření konců kolon • Vysoká účinnost vyrobených kolon Apparatus for filling of packed capillary columns 1 - CO2 cylinder 2 - HPLC pump 3 - manometer 4 - on/off valve, 5 - stainless steel filling reservoir, 6 - fused silica capillary, 7 - restrictor Examples of SFC separations SFC separation of Rudolf II seal (beeswax). Column 320 pm x 150 mm, 5 pm Biospher C18, t=80°C, FID 150°C, program 8-35 MPa Examples of SFC separations J SFC separation of poly(dimethylsiloxane). Column 320 um x 150 mm, 5 urn Biospher C18, t=80°C, FID 150°C, pressure program 8-35 MPa SFC today ? - chiral separations (enantiomers) - separation and purification in pharmaceutical research/industry 1 higher throughput than HPLC (more separations/analyses per unit time puro COa -> CQ2 + MeOH SFC as 2nd dimension in 2D chromatographic separations Ionic Liquids (ILs) ? = liquid organic salts (melting point below 100°C) = liquids composed exclusively of ions (no electroneutral particles) properties of ILs differ markedly from those of common molecular solvents (water, organic solvents) number of ..possible" ILs = ~1018 CH3 Ionic liquids - anions J F F History of Ionic Liquids ...1914...Walden... [(C2H5)4N][N03] • 1980s: Chloroaluminate Ionic Liquids 1st generation \—/' [AICIJ- J.S. Wilkes, J.A. Levisky, R.A. Wilson and C.L. Hussey, Inorg. Chem. 21 (1982) 1263-1264. • 1990s: Air- and moisture-stable Ionic Liquids 2nd generation ^ ^ J X=f [BFJ- J.S. Wilkes and M.J. Zaworotko, J. Chem. Soc. Chem. Commuh (1992) 965-966. • 2000s: First examples of ..Task Specific Ionic Liquids" 3rd generation \_ ,-s-\ \=J [PFJ A.E. Visser, R.P. Swatloski, W.M. Reichert, R. Mayton, S. Sheff, A. Wierzbicki, J.H. Davis, Jr. and R.D. Rogers, Chem. Commuh. (2001) 135-136. • 2010s: Biodegradable Ionic Liquids 4th generation IL H3C jN-C6H13 F O OF I II - II I F—C-S—N-S-C-F I II II I F O OF [hmim][Tf2N] I siloxane polymer (PDMS, M„ ~ 4.8x105) open tubular capillary column -micropacked dmeeb1 ■■■■ CAUSES — SEIFTE MRUS >r» ID SKIN MB §'fS Relative partition coefficients in [bmim][BF4]-scC02 system A A 1 0 0 <> O ♦ ♦ ♦ * m 1 V V a fl 300 400 500 600 700 800 Pm I kg.rrr3 O aniline • anisole □ azulene ■ benzoic acid A 1-hexanol A indole V W,W-dime1hylaniline ▼ W-melhylaniline O p-cresol ♦ phenelhyl alcohol @ phenol B: naphthalene Review papers on SFC applications a) pharmaceutical analysis E. Lemasson, S. Bertin, C. West: Use and practice of achiral and chiral supercritical fluid chromatography in pharmaceutical analysis and purification, /. Separ. Set 2016, 39, 212-233; 62. V. Desfontaine, D. Guillarme, E. Francotte, L. Nováková: Supercritical fluid chromatography in pharmaceutical analysis, /. Pharm. Biomed. Anal. 2015, 113, 56-71; J. M. Plotka, M. Biziuk, C. Morrison, J. Namiesnik: Pharmaceutical and forensic drug applications of chiral supercritical fluid chromatography, TrAC - Trends Anal. Chem. 2014, 56, 74-89; htl 12 . b) food analysis J. L. Bernal, M. T. Martin, L. Toribio: Supercritical fluid chromatography in food analysis, /. Chromatogr. A 2013,1313, 24-36; htl 22. 2) SFE - Supercritical fluid extraction (C02) solid samples aqueous samples solvent power of C02 depends strongly on pressure (density) ? chrom. analysis - decompression - analyte losses - trapping ? Basic technique of analyte trapping into organic solvent + simple construction - high consumption of solvent ■ loss of volatile analytes ■ restrictor clogging SFE of (pleasing) liquid samples ? Wine analysis ä.. <5 1 1 Direct Continuous Supercritical Fluid Extraction as a Novel Metho of Wine Analysis: Comparison with Conventional Indirect Extraction and Implications for Wine Variety Identification d Pavel Karásek, Josef Planeta, Elena Varaďová Ostrá, Milena Mikešová, Jan Goliáš, Michal Roth, and Jiří Vejrosta Journal of Ch romatography A 2003, J002, 13-23. | 0 1 vuSSkT l ; 8 y l_i L 1 y m brnenská velkopavlqvická ■ ■ k mikulovská =cdluží u 5ř AURELUS FRANKOVKA f* ténkká enecká efsk0-hradišťská Miiller Thurgau MIISKATOTl'U.NEL f# IVEIIBURGSKE PALAVA 1* pOIITIIgalské modré "*>~ í f* Kiilandské bílé HILANDSKÉ MODRÉ f* RULA1VDSKÉ ŠEDÉ Ky/link vlašský Smivignon f* SVATOVAVRIfVECKÉ f* Veltlínské čtrvení f* - ■ :. - - 121 wine samples 21 grape varieties (Vitis vinifera L.) 4 vintages (years 1996-1999) ZWEKiliLTREBE f* Multivariate Statistics of the wine varieties represented by >~4 wine samples cluster analvsisl - used to select the 4 samples/variety if more discriminant analysis - elimination of redundant (= linearly dependent) component peak areas from the input data matrix canonical correlation analysis - computation of discriminant functions, i.e., the latent factors differentiating among the wine samples Info: http://www.statsoft.com/textbook/stathome.html Computation tool: KyPlot spreadsheet SW, Koichi Yoshioka, http://www.qualest.co.jp/Download/KyPlot/kyplot_e.htm, http://www.kyenslab.com/en Statistical processing (discriminant analysis) of chromatograms Compared with the procedure involving solid phase extraction (SPE-SEE-GC), direct SFE of wines (DCSFE-GC) provides much clearer discrimination among wine varieties 9 If 8 DCSFE-GC SPE-SFE-GC Other analytical applications of direct continuous SFE (DCSFE) of aqueous samples: •beer - aldehydes, ketones, fatty acids, esters; PAH, PCB (DCSFE-GC) •natural insecticides - Pyrethrines {Chrysanthemum cinerariaefoliiim) (DCSFE-HPLC) •lycopene (tetraterpene, red dye of tomatoes, etc.) (DCSFE-HPLC) X x. Y J Interphase distribution of analytes in H20 - scC02 system Partition Coefficient [K = ylx) of Salicylic Acid as a Function of rand P C02-Water Partition Coefficients (333 K, 20 MPa) vs. Octanol-Water Partition Coefficients at Ambient Conditions Red symbols = otffio-substltuted phenols Blue symbol = 2 dissociable protons Review articles on SFE applications J. A. Mendiola, M. Herrero, A. Cifuentes, E. Ibanez: Use of compressed fluids for sample preparation: Food applications, J. Chromatogr. A 2007,1152, 234-246; 46. M. Herrero, J. A. Mendiola, A. Cifuentes, E. Ibanez: Supercritical fluid extraction: Recent advances and applications, J. Chromatogr. A 2010,1217, 2495-2511; http://dx.d0i.0rg/l 0.1016/j .chroma.2009.12.019. C. G. Pereira, M. A. A. Meireles: Supercritical Fluid Extraction of Bioactive Compounds: Fundamentals, Applications and Economic Perspectives, Food Bioprocess. Technol. 2010, 3, 340-372; b2 . J. Azmir, I. S. M. Zaidul, M. M. Rahman, K. M. Sharif, A. Mohamed, F. Sahena, M. H. A. Jakurul, K. Ghafoor, N. A. N. Norulaini, A. K. M. Omar: Techniques for extraction of bioactive compounds from plant materials: A review, J. Food Eng. 2013, 11 7, 426436; htl 14. M. M. R. de Melo, A. J. D. Silvestře, C. M. Silva: Supercritical fluid extraction of vegetable matrices: Applications, trends and future perspectives of a convincing green technology, J. Supercrit. Fluids 2014, 92, 115-176; http://dx.d0i.0rg/l0.1016/i.supflu.2014.04.007 . A. R. C. Morais, A. M. D. Lopes, R. Bogel-Lukasik: Carbon Dioxide in Biomass Processing: Contributions to the Green Biorefinery Concept, Chem. Rev. 2015,115, 3-27; http://dx.doi.org/10.102 l/cr500330z . 3) Extraction with liquids at elevated temperatures [T> Tboi|solvent] and pressures [P>Psatsolvent(7)] organic solvents / blends: PFE, PLE, PSE, ASE PFE instruments - automated extractors: Advantages of PFE compared with low-pressure Soxhlet extrc a) higher solubility of analytes because of their higher volatility at elevated temperature b) | fialytes from the sa____ Ire (faster masstra factions) Ind b) results in fas lower may be tune Lch less than in SF of organic solvent: ivironment inT f) tighter control of composition when using mixed solvents -unlike the Soxhlet extraction, PFE does not involve any solvent phase transition (vapor-liquid equilibrium) PFE application - nutrition-relevant substances from plants 1) hops (cones, pellets) alpha acids (humulones) beta acids (lupulones) isohumulones R - -CH(CH3)2, -CH2CH(CH3)2, -CH(CH3)CH2CH3 2) "Tea" plants honeybush (Cyclopia intermedia) - South Africa rooibos (Aspalathus linearis) - South Africa čaj (Camelia sinensis) - China yerba mate (Ilex paraguayensis) - South America Review papers on PFE applications a) Food analysis: A. Mustafa, C. Turner: Pressurized liquid extraction as a green approach in food and herbal plants extraction: A review, Anal. Chim. Acta 2011, 703, 8-18; http://dx.doi.Org/l0.1016/i.aca.2011.07.018 . A. Baiano: Recovery of Biomolecules from Food Wastes - A Review, Molecules 2014,19, 14821-14842; http://dx.d0i.0rg/l 0.3390/molecules 190914821 . C. C. Teo: Pressurized hot water extraction (PHWE), J. Chromatogr. A 2010,1217, 2484-2494; http://dx.doi.org/10.1016/i.chroma.2009.12.050 . S. M. Zakaria, S. M. M. Kamal: Subcritical Water Extraction of Bioactive Compounds from Plants and Algae: Applications in Pharmaceutical and Food Ingredients, Food Eng. Rev. 2016, 8, 23-34; Lx. 4) Pressurized hot (subcritical) water extraction Motivation: Water is not only the greenest but also the most tuneable solvent (through changes in operating Tand P). Standard conditions (25 °C, 0.1 MPa): NaCI well soluble, benzene nearly insoluble "Supercritical" conditions (>374 °C, >22.1 MPa): NaCI ~ insoluble, benzene ~ fully miscible Applications of high temperature, high pressure water: a) Supercritical water (f > 374 °C, P > 22 MPa) supercritical water oxidation, SCWO supercritical water dissolves Si02 - geochemistry, surface: b) Subkritickavoda(100 °C < f<374°C, P>Ps*{t)) „environmental remediation" extraction of plant materials analytical chemistry - sample preparation biopolymers- cellulose dissolution, protein hydrolysis biomass gasification - energy (CO+H2) Motivation water = the "greenest" and the most "tuneable" solvent Property "ambient" 25 °C, 0.1 MPa Density pi kg-rrf3 997.0 Cohesive enerqy density cl J-crrr3 2299 -— -- Solubility parameter SI (J-crtr3)"2 47.9 Internal pressure Pini / MPa 169 Ion product K,l (mol-drrT3)2 1x10" Relative permittivity e, 78.4-- -- PHWE : 100°C < f< 374°C, P> Psat (f) relative wealth of analytical applications of PHWE X relative lack of solubility data PHW: "environmental" prediction naphtha Ig no anthracGriG pyrGnG chrysGnG 1,2-bGnzanthracGnG triphGnylGriG pGrylGnG p-tGrphGnyl fluorGnG fluoranthGnG naphthalGriG, Gnvi biphenyl, Gnvi acGnaphthGnG, Gnvi fluorGnG, Gnvi anthracGnG, Gnvi phGnanthrGnG, Gnvi pyrGnG, Gnvi fluoranthGnG, Gnvi chrysGnG, Gnvi 1,2-bGnzanthracGne triphonylGnG, Gnvi naphthacGno, Gnvi bGnzo[a]pyrGnG, Gnvi dibGnz[a,h]anthracGnG, Gnvi p Gry Ig no, Gnvi bGnzo[g,h,i]pGrylGnG, Gnvi corononG, Gnvi 0.14 mg/m3(25°C, 0.1 MPa) Smartphones, tablets, etc. ... OLED displays Increasing production => increasing rate of disposal Environment ?? ... Aqueous solubilities l,3,5-tris(diphenylamino)benzene 4,4'-bis(N-carbazolyl)-l,l'-biphenyl 9,10-bis(phenylethynyl)anthracene Aqueous solubilities: diamondoids vs. PAHs Group contributions to log(activity coefficient) Raoult-law ideality ^^&Z\^JfB^™_^ hydrophobic 1 k~ 0 -1 o CH(arom) • C(arom) ■ CH2(ring) ■ N(pyridine) A S(ring) A O(ring) V NH(pyrrole) T C=0 + CH(aliph) O N(f-amine) z , h.ydrophilic 300 350 400 450 500 Tl K Activity coefficients: diamondoids vs. PAHs Tapered capillaries 1 cm 1 cm 1 cm J cm .. i i i i 48 cm T70fim ■V Tapered capillaries - internal diameter profiles 90 mg/min 0 10 20 30 40 50 60 70 80 90 100 capillary length [cm ] Tapered capillaries - what are they good for ? Karel Slais (1995) - theory - tapered capillaries can improve resolution of ampholytes in capillary isoelectric focusing (CIEF) 90 mg/min 260 mg/min 360 mg/min 500 mg/min 20 30 40 50 60 70 80 90 100 capillary length [cm ] Tapered capillaries - how do they work in CIEF ? conventional cylindrical capillary tapered capillary A, __3 Separation of several Dickeya bacterium species with mutually close isoelectric points by CIEF with 2.0-4.0 pH gradient. Left - conventional cylindrical capillary; Right - tapered capillary. Inner surface-etched capillaries of uniform diameter Etching with SCW can also be arranged so that the capillary internal diameter remains constant but the inner surface acquires a "porous" structure" with cavities of defined size MRSA = methicillin-resistant Staphylococcus aureus MSSA = methicillin-sensitive Staphylococcus aureus Columns - troubles in surface treatment - an example > untreated fused silica capillary (100 umi.d.) > fused silica capillary after etching with 2-chloro-l,l,2-trifluoroethyl methyl ether (33 % of capillary volume filled with the liquid ether, capillary sealed, 320 °C, 12 hours) followed by etching with saturated methanolic solution of ammonium hydrogen difluoride (25 °C, 24 hours) > fused silica capillary after etching with 2-chloro-l,l,2-trifluoroethyl methyl ether (50 % of capillary volume filled with the liquid ether, capillary sealed, 350 °C, 12 hours); black coloration comes from the carbon soot produced by decomposition of the ether Separation of Nucleic Acid Bases and their Derivatives Thank you for your attention Comparison of isocratic elution on bare silica monolithic (A) and sulfoalkylbetaine monolithic (B) capillary cmr«niin ;r.-mr.lr. -,,1-4*. 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