[ viridis columns ]

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viridis Columns

Bringing a New Dimension to SFC

Combining state-of-the-art media manufacturing

with industry leading column technology, Viridis®

Supercritical Fluid Chromatography (SFC) Columns

bring a new level of reproducibility to the world of

laboratory-scale purification.

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BEH 2-Ethylpyridine

BEH

CSH Fluoro-Phenyl

Silica 2-Ethylpyridine

Silica

Manufactured for Reliability and Consistency

Based upon over 50 years of experience in the separation

science industry, Waters chromatographic media sets the

standard for quality and batch-to-batch reproducibility.

As a primary manufacturer of silica and hybrid

chromatographic particles, all stages of product synthesis

are meticulously monitored and controlled, ensuring

complete characterization of the finished product.

This level of manufacturing process understanding

delivers chromatographic media with guaranteed

consistent performance year after year.

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Viridis Hybrid Columns

Based on the patented Ethylene-Bridged Hybrid (BEH) particle technology and Charged-Surface Hybrid (CSH™) particle technology, Viridis Hybrid Columns offer a wide range of selectivities and batch-to-batch reproducibility. The reduction and control of surface silanol activity on Viridis Hybrid particles delivers, under SFC conditions, excellent peak shapes—without the use of mobile-phase additives—even for well-retained basic achiral compounds.

Three available surface chemistries extend the selectivity range of SFC separations:

■ BEH 2-EtHylpyridinE (2-Ep)

Exhibits good retention, peak shape and selectivity properties without the use of additives.

■ BEH

Heightens interaction with polar analyte functional groups, when compared to the less polar surface of BEH 2-EP.

■ CSH Fluoro-pHEnyl

Provides good retention for weak bases and alternate elution orders for acidic and neutral compounds, via significantly different selectivity.

ChemistryparticleShape

particle Size (μm)

pore Size(Å)

SurfaceArea

(m2/g)

pore Volume (cc/g)

Carbonload (%)

Endcapped

BEH 2-Ep Spherical 5 135 185 0.70 9 no

BEH Spherical 5 135 185 0.70 n/a n/a

CSH Fluoro-phenyl

Spherical 5 135 185 0.70 10 no

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0 9.5 10.0 min

Viridis CSH Fluoro-PhenylS = 72Part Number: 186005694

Viridis BEHS = 56Part Number: 186005725

Viridis BEH 2-EPPart Number: 186005756

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1

3 4

5

6 7 8

1

1

2

2

2

3

3

4

4

5

5 6

7

7

8

8

Viridis Hybrid SFC Columns are engineered to provide a wider range of selectivity options.

Columns: 4.6 x 150 mm, 5 µm Gradient: 8 minutes Mobile Phase: 5–40% methanol without additive Flow Rate: 3.0 mL/min Column Temp.: 40 °C Backpressure: 2175 psi (150 bar) Injection Volume: 3 µL

Compounds:1. Coumarin2. Flavone3. Caffeine4. Thymine5. Papaverine6. Sulfamethoxazole7. Cytosine8. Sulfamethizole

Enhanced Selectivity

Selectivity Values

Viridis Hybrid Columns are designed to offer a wide selectivity range. One measure of how different (or similar) the selectivities of two column chemistries are under a given set of SFC conditions, is the Neue Selectivity Value (S).* As shown below, this parameter may be calculated from a plot of retention factors (k) or retention times (tR) for analytes on a pair of columns under the same chromatographic conditions. A higher selectivity value indicates a higher degree of orthogonality between two columns and/or two sets of chromatographic conditions.

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tR Viridis BEH 2-Ethylpyridine

tR Vi

ridis

CSH

Flu

oro

Phen

yl

0

1

2

3

4

5

6

0 1 2 3 4 5 6 7

R2 = 0.480

S = 72

The Neue Selectivity Values may be used to estimate the degree of orthogonality between the selectivities of a pair of columns under the same LC or SFC conditions. It is calculated as shown from the coefficient of determination (R2 = 0.480) of a plot of retention times (or retention factors).* As seen below, on a scale of 0 to 100, a higher S value indicates a higher degree of orthogonality. Compounds: caffeine, theophylline, ibuprofen, fenoprofen, hydrocortisone, ketoprofen, sulfamethoxazole, sulfamethizole, coumarin, flavone, adenine, thymine, uracil, prednisone, prednisolone.

Selectivity Values (S) compared for pairs of Viridis SFC Columns. S may range from 0 (identical selectivity) to 100 (maximum selectivity difference or orthogonality).* Viridis Hybrid SFC columns are shown in light blue.

Silica 2-Ep

0

Silica 78 0

CSH Fluoro-phenyl

85 38 0

BEH 75 11 41 0

BEH 2-Ep

25 60 72 56 0

Silica 2-Ep

SilicaCSH

Fluoro-phenyl

BEHBEH 2-Ep

* (1) U.D. Neue, E.S. Grumbach, J.R. Mazzeo, K. Tran, and D.M. Wagrowski-Diehl “Method development in reversed-phase chromatography” Chap. 6 in: I.D. Wilson, ed. Bioanalytical Separations, Handbook of Analytical Separations, Vol. 4 Elsevier, Amsterdam (2003); (2) U.D. Neue, J.E. O’Gara, and A. Méndez “Selectivity in reversed-phase separations: influence of the stationary phase” J. Chromatogr. A 1127(1–2): 161–174 (2006); (3) U.D. Neue and A. Méndez “Selectivity in reversed-phase separations: general influence of solvent type and mobile phase pH” J. Sep. Sci. 30(7): 949–963 (2007).

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Viridis Silica Columns

Based on Waters long history of chromatographic silica production, the Viridis Silica Columns are designed to be highly reproducible and predictable based on tight product specifications and very low metal content. They are available for both analytical screening and in preparative column dimensions for purification. Separation methods can be optimized and scaled up to Viridis Prep OBD™ Columns.

■ SiliCA 2-EtHylpyridinE (2-Ep)

Widely used in achiral SFC separations exhibiting good retention, peak shape and selectivity properties both with and without the use of additives.

■ SiliCA

Unbonded silica heightens interaction with polar compound functional groups, when compared to the less polar surface of Silica 2-EP column chemistry, resulting in different selectivity.

ChemistryparticleShape

particle Size (μm)

pore Size(Å)

SurfaceArea

(m2/g)

pore Volume (cc/g)

Carbonload (%)

Endcapped

Silica 2-Ep Spherical 5 100 340 0.90 8 no

Silica Spherical 5 100 340 0.90 n/a n/a

High Mass loading

1 2

3

4

0.00 0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75 3.00 min

Column: Viridis Silica 2-EP OBD, 19 x 150 mm, 5 µmPart Number: 186004945Gradient Separation: 4.2 minutesMobile Phase: 5–15% methanol with no additiveFlow Rate: 85 g/min at 40 °CSample Conc.: 80 mg/mL Wavelength: 220 nmSystem Backpressure: 120 bar

Injection Vol.: 750 µL Total Mass Load: 60 mg

Injection Vol.: 350 µLTotal Mass Load: 28 mg

Injection Vol.: 100 µLTotal Mass Load: 8 mg

Compounds: 1. Coumarin2. Flavone3. Ibuprofen4. Carbamazepine

Viridis Silica 2-EP Columns provide excellent mass loading to ensure maximum throughput.

Viridis Silica Column vs. Viridis Silica 2-Ep Column

1 2

3

2

1

3

0.0

2.0e-1

4.0e-1

6.0e-1

0.0

2.0e-1

4.0e-1

6.0e-1

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 min

Gradient Separation: 4.2 minutesMobile Phase: 5–15% methanol with no additiveFlow Rate: 85 g/min at 40 °CWavelength: 220 nmSystem Backpressure: 120 bar

Compounds:1. Flavone2. Ibuprofen3. Carbamazepine

Using the same SFC gradient conditions, different selectivity can be achieved by using a Viridis Silica Column or Viridis Silica 2-EP Column. The retention time for ibuprofen was reduced and the carbamazepine retention time was increased by switching column chemistries, with little change in the retention of flavone.

Viridis Silica 2-EP OBD Column19 x 150 mm, 5 µmPart Number: 186004945

Viridis Silica OBD Column19 x 150 mm, 5 µmPart Number: 186004918

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OBD Column Design—Improved Mechanical Stability

Laboratory-scale SFC purification presents many challenges to the chromatographer. Inconsistency in column-to-column performance and lifetimes often result in lost samples, repeat purification runs and poor scalability from small- to large-volume columns. Common SFC preparative column failures are due to mechanical instabilities, especially voids in the packed bed. The Viridis Preparative OBD™ Columns combine the patented* optimized packing process with a unique set of column designs ensuring increased lifetime and greater reproducibility. All Viridis Column characteristics are available in the preparative OBD column format.

* US Patent # 7,399,410, UK Patent # GB2408469

Batch-to-Batch reproducibility

Waters is dedicated to maintain the tightest specifications in the SFC industry. Controlled manufacturing processes and column packing procedures ensure that you receive the best, most reproducible SFC column available.

1 2

3 4

5 6

0 1 2 3 4 5 6 7 8 9 10 11 min

Batch A

Batch B

Batch C

Batch D

Batch E

5 µm

This excellent reproducibility is a result of our commitment to maintaining the tightest specifications in the industry. Viridis Hybrid Columns start with high purity raw materials, and are produced using controlled manufacturing processes and column packing procedures that provide today’s scientists with the most reproducible SFC column available.

Column: Viridis BEH 2-EP, 2.1 x 150 mm, 5 µmPart Number: 186006545Flow Rate: 0.65 mL/min Mobile Phase: 90/10 carbon dioxide/methanolBackpressure (BPR): 2175 psi (150 bar)Column Temp.: 40 °CSystem: UPSFC (alpha)Injection Vol.: 1.0 μLDetection: UV at 220 nm (compensated wavelengths 350–450 nm)

Compounds:1. Toluene 2. Flavone 3. Thymine 4. Papaverine 5. Prednisolone 6. Sulfanilamide

Ordering Information

Viridis Analytical SFC Columns

dimensionparticle

SizeBEH 2-Ep BEH

CSH Fluoro-phenyl

Silica 2-Ep Silica

2.1 x 150 mm 5 µm 186006545 186006544 186006543 186006542 186006541

3.0 x 50 mm 5 μm 186005750 186005719 186005688 186005800 186005804

3.0 x 100 mm 5 μm 186005751 186005720 186005689 186005801 186005805

3.0 x 150 mm 5 μm 186005752 186005721 186005690 186005802 186005806

3.0 x 250 mm 5 μm 186005753 186005722 186005691 186005803 186005807

4.6 x 50 mm 5 μm 186005754 186005723 186005692 186004935 186004908

4.6 x 100 mm 5 μm 186005755 186005724 186005693 186004936 186004909

4.6 x 150 mm 5 μm 186005756 186005725 186005694 186004937 186004910

4.6 x 250 mm 5 μm 186005757 186005726 186005695 186004938 186004911

Viridis preparative SFC Columns

dimensionparticle

SizeBEH 2-Ep BEH

CSH Fluoro-phenyl

Silica 2-Ep Silica

10 x 50 mm 5 μm 186005758 186005727 186005696 186004939 186004912

10 x 100 mm 5 μm 186005759 186005728 186005697 186004940 186004913

10 x 150 mm 5 μm 186005760 186005729 186005698 186004941 186004914

10 x 250 mm 5 μm 186005761 186005730 186005699 186004942 186004915

OBD 19 x 50 mm 5 μm 186005762 186005731 186005700 186004943 186004916

OBD 19 x 100 mm 5 μm 186005763 186005732 186005701 186004944 186004917

OBD 19 x 150 mm 5 μm 186005764 186005733 186005702 186004945 186004918

OBD 19 x 250 mm 5 μm 186005765 186005734 186005703 186004946 186004919

OBD 30 x 50 mm 5 μm 186005766 186005735 186005704 186004947 186004920

OBD 30 x 75 mm 5 μm 186005767 186005736 186005705 186004948 186004921

OBD 30 x 100 mm 5 μm 186005768 186005737 186005706 186004949 186004922

OBD 30 x 150 mm 5 μm 186005769 186005738 186005707 186004950 186004923

OBD 30 x 250 mm 5 μm 186005770 186005739 186005708 186004951 186004924

OBD 50 x 50 mm 5 μm 186005771 186005740 186005709 186004952 186004925

OBD 50 x 100 mm 5 μm 186005772 186005741 186005710 186004953 186004926

OBD 50 x 150 mm 5 μm 186005773 186005742 186005711 186004954 186004927

OBD 50 x 250 mm 5 μm 186005774 186005743 186005712 186004955 186004928

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www.waters.com

Waters and Viridis are registered trademarks of Waters Corporation. BEH Technology, CSH, OBD, UPSFC, and The Science of What’s Possible are trademarks of Waters Corporation.

All other trademarks are the property of their respective owners.

©2012 Waters Corporation. Printed in the U.S.A. July 2012 720003326EN SC-FP

www.waters.com/viridis

The quality management system of Waters’ manufacturing facilitiesin Taunton, Massachusetts and Wexford, Ireland complies with the International Standard ISO 9001 Quality Management and Quality Assurance Standards. Waters’ quality management system is periodically audited by the registering body to ensure compliance.