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Comparison of CCS Instruments

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The different types of instruments are compared in terms of capacity (column volume), resolution, and duration required for the separation of targeted compounds.
This workshop presentation was prepared by Dr. Friesen (http://www.dom.edu/departments/physicalsciences/faculty/j-brent-friesen).

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Comparison of CCS Instruments

  1. 1. CCS Instrument Comparison https://upload.wikimedia.org/wikipedia/commons/3/39/Music_wall.jpg
  2. 2. Liquid/Liquid Separation (LLS) Kuhni Extraction Columns Continuous Mixer-Settler Liquid-Liquid Extraction Liquid/Liquid Chromatography A. Martin & R. Synge Centrifugal Countercurrent Separation (CCCS) Gravitational Countercurrent Separation (GCCS) Countercurrent Chromatography (CCC) “hydrodynamic” Centrifugal Partition Chromatography (CPC) “hydrostatic” Droplet Counter Current Chromatogr. Craig Counter Current Distribution Kostanyan Pulsed Rotational Locular Separatory Funnels Countercurrent Separation (CCS) Friesen JB, McAlpine JB, Chen SN, Pauli GF Countercurrent Separation of Natural Products: An Update Journal of Natural Products 78: 1765-1796 (2015) dx.doi.org/10.1021/np501065h
  3. 3. CCC vs CPC Berthod 2007 J of Liq Chromatog & Relat Technol volume 30 p.447
  4. 4. Survey Results instrument volume articles TBE-300A, TBE- 300B 260-300 105 TBE-20A 16 2 TBE-1000A 1000 10 TBE-5000A 5000 1 GS-20A2 40 11 GS-10AB, GS10A- 2, GS10A 230-240 54 Spectrum 15 mini 9 midi 15 maxi 2 FCPC 50, 200, 1000, 5000 30 CCC-1000 300-325, 850 29 PC-Inc. 8 instrument volume articles HPCPC LLB-M 8 Sanki CPC 80, 240, 1400, 5400 6 Custom 400 - 15000 20 DCCC 3 Armen TC TC-250 1 SPOT CPC 100 light 2 HHS-400A ? 3 custom CPC 1 EMC-500A ? 1 IFBE custom 420 7 Quattro 4 D400, D1200 ? 2 SRCCC 1 Friesen 2015 Journal of Natural Products vol 78 p. 1764
  5. 5. Column Volumes 24 34 78 104 17 12 49 13 16-100 108-200 220-250 260-300 320-400 420-500 600-1000 1200-22000 Column Volumes: All CS Instruments number of instruments columnvolumes(mL) Friesen 2015 Journal of Natural Products vol 78 p. 1764
  6. 6. GUESSmix in hexane / ethyl acetate / methanol / water 4:6:4:6 0 0.25 0.5 0.75 1 1.25 1.5 1.75 2 2.29 2.67 3.2 4 5.33 8 16 ¥K'(2) A 0 0.25 0.5 0.75 1 1.25 1.5 1.75 2 2.29 2.67 3.2 4 5.33 8 16 ¥K'(2) A 280nm 230nm J-type centrifuge 120 mL Fast Centrifugal Partition Chromatography (FCPC) 200 mL 220 mg 440 mg G H X T r C D F R U V A Q M N Z E O I Y b
  7. 7. Figure 4. CCC and CPC chromatograms obtained by using CUAE/ CCC/CPC system. (a) CCC chromatogram using solvent system of HEMWat at a volume ratio of 4:3:4:3; (b) (b) CPC chromatogram using solvent system of HEMWat at a volume ratio of 4:3:4:3; Development of Circulating Ultrasounic-Assisted Online Extraction Coupled to Countercurrent Chromatography and Centrifugal Partition Chromatography for Simultaneous Extraction and Isolation of Phytochemicals: Application to Ligusticum chuanxiong Hort Yuchi Zhang, Chunming Liu, Yanjuan Qi, Yuchun Li, and Sainan Li Ind. Eng. Chem. Res., 2015, 54 (11), pp 3009–3017 DOI: 10.1021/ie504179r TAUTO TBE−300B SIC CPC-240 (Kyoto, Japan) instrument that was modified in our laboratory Comparison of HSCCC & CPC
  8. 8. Figure 4. CCC and CPC chromatograms obtained by using CUAE/ CCC/CPC system. (c) CCC chromatogram using solvent system of HEMWat at a volume ratio of 4:3:4:2; and (d) CPC chromatogram using solvent system of HEMWat at a volume ratio of 4:3:4:2. Development of Circulating Ultrasounic-Assisted Online Extraction Coupled to Countercurrent Chromatography and Centrifugal Partition Chromatography for Simultaneous Extraction and Isolation of Phytochemicals: Application to Ligusticum chuanxiong Hort Yuchi Zhang, Chunming Liu, Yanjuan Qi, Yuchun Li, and Sainan Li Ind. Eng. Chem. Res., 2015, 54 (11), pp 3009–3017 DOI: 10.1021/ie504179r Comparison of HSCCC & CPC
  9. 9. Fig. 4. CCC, CPC of UADE extract of white peony roots and fractions. (a) CCC chromatogram and (b) CPC chromatogram obtained using the EBuEtWat (1:3.5:2:4.5, v/v/v/v) solvent system. (c) UHPLC chromatogram of the UADE extract of white peony roots. (1) Albiflorin, (2) benzoylpaeoniflorin, (3) paeoniflorin, and (4) galloylpaeoniflorin. Ultrason Sonochem. 2015 26:111-118. doi: 10.1016/j.ultsonch.2015.02.017. Development of "ultrasound-assisted dynamic extraction" and its combination with CCC and CPC for simultaneous extraction and isolation of phytochemicals. Zhang Y, Liu C, Li J, Qi Y, Li Y, Li S. Comparison of HSCCC & CPC
  10. 10. Fig. 1. Solid line represents reversed-phase separation of standards using Pharma-Tech CCC-1000, 320mL column volume, Sf = 68%, HEMWat 4:6:4:6 1.5m L/min, loading 1–5 mg of each 12 standards (Aspirin (A), caffeine (C), nicotinic acid (D), ferulic acid (F), red new Coccine (R), umbelliferone (U), vanillin (V)). Dashed line is the separation of the same mixture using DE Spectrum semi-preparative coils at 54×g (751 rpm), 136mL column volume, lower layer as mobile phase, 1.5mL/min, head to tail elution, Sf =73 %, 60mg total loading (5mg of each 12 standards as above), monitored by DAD 254 nm, compared separation time = 130 min. Journal of Chromatography A, 1216 (2009) 4181–4186 Performance comparison using the GUESS mixture to evaluate counter-current chromatography instruments. Hacer Guzlek, Philip LeslieWood, Lee Janaway Comparison of HSCCC & HPCCC
  11. 11. Fig. 4. Separation of caffeine and vanillin with the Arizona L heptane–ethyl acetate–methanol–water 2:3:2:3 (v/v) liquid system. Reversed-phase mode: organic upper stationary phase and aqueous lower mobile phase flown in the head to tail direction. Top chromatograms: column HPCCC Mini 19.6mL with a 0.8mm I.D. tubing coil, 2100 rpm, 2 mL/min. Sf = 38%, Rs = 2.0. Bottom chromatograms: column HPCCC Mini 20.8mL with a 1.6mm I.D. tubing coil, 1800 rpm, 5mL/min, Sf = 66%, Rs = 2.0. Injectionvolume 0.5 mL, concentration 1mg/mL in lower mobile phase. Detection UV at 254 nm. Left chromatograms shown in volume units. Right chromatograms shown in time units. Berthod 2009 Journal of Chromatog A vol 1216 p 4169 HPCCC Mini 19.6mL 0.8 mm tubing HPCCC Mini 20.8mL 1.6 mm tubing Comparison of HPCCC designs
  12. 12. Fig. 5. Separation of nine GUESS solutes with HepEMWat 1:1:1:1 Organic upper stationary phase. Top chromatograms obtained in 22min: column HPCCC Mini 19.6mL with a 0.8mmI.D. tubing coil, 2100 rpm, 2 mL/min. Sf = 46%. Bottom chromatograms obtained in 13min: column HPCCC Mini 20.8mL with a 1.6mm I.D. tubing coil,1800 rpm, 5 mL/min, Sf = 72%. Injection volume 0.5 mL, concentration ∼1mg/mL (each) in lower mobile phase. Detection UV at 254 nm. The inset chromatograms are enlargements of the early eluting peaks.. Carvone (O) retention time is indicated. HPCCC Mini 19.6mL with 0.8mmI.D. tubing coil HPCCC Mini 20.8mL 1.6mm I.D. tubing Berthod 2009 Journal of Chromatog A vol 1216 p 4169 Comparison of HPCCC designs

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