Agilent Prep LC Columns High Performance Preparative Columns for Maximum Sample Purification Manual
Contents
1. www agilent com chem Agilent Prep C18 30 x 250 mm 10 um p n 410910 302 Agilent Technologies2. Stability Increased throughput requires improved prepara tive column stability for reliable and reproducible results Agilent Prep columns demonstrate excel lent stability for more than 1000 injections under a variety of solvent conditions This is demonstrated using the Agilent Prep C18 21 2 mm x 150 mm 10 um column in Figure 4 This allows for increased productivity by providing the stability to run unattended for long periods of time Column Mechanical Stability for 1000 Injections Agilent Prep C18 21 2 mm x 150 mm 10 pm 9000 2 0 8000 NN ce ca a is 7000 4 p18 N Efficiency tee TF5 Tailing factor at 6000 m17 5 of peak height 4 4CINB 4 Chloronitrobenzene 2 16 2 5000 4 g S H15 Phenol N 4000 4 4CINBN wi 14 gt o Toluene N aoo ea e Phenol TF5 0 2000 fs ee re P a po a leas o 4CINB TF5 x fey 7 ane 4 s 4 e Toluene TF5 1000 L11 0 i 1 0 1 2 3 4 5 6 7 9 10 Injection number 100 No changes in efficiency and tailing over 1000 injections indicates the column is stable and the packed bed is not compressing Figure 4 Agilent Prep C18 column mechanical stability over 1000 injections using many solvent types Flexibility for All Your Purification Needs preparative columns are available in a wide range of dimensions to meet all of your preparative Whether it is improving loadability by using high needs See Tables 3 and 4 Ag
3. compounds Table 1 compares the loadability of methyl and ethyl benzoate on an Agilent Prep C18 column against two competitors Table 1 Loadability Comparison mg loadedon mg loaded mg loadable in Column at Rs 1 25 per g packing 50 x 250 mm column Agilent Prep C18 4 80 3 47 1735 Competitor K C18 4 45 2 68 1340 Competitor W C18 3 88 2 32 1160 Mobile phases A H 0 B Methanol isocratic 70 5 78 B initial resolution 1 8 Column temperature 30 C Injection 125 uL of each dilution Sample 100 mg mL each of methyl benzoate and ethyl benzoate in 90 methanol This unique bonded phase also provides outstand ing peak shape with basic compounds allowing very high purity sample isolation See Figure 2 Agilent Prep C18 Competitor W C18 mAU J mAU 4 J a J ae a 4000 J J J i i J 3500 i 3500 5 4 i j i J J 1h 7 ni i l j geig ao a157 2500 a 2500 4 if all ii 2000 4 eA 2000 J 11h J 4 J 19007 ATAN C 50 mg mL 15005 50 mg mL 7 ON a J q z 7 1000 5 e SEE Ae ne EAE 1000 a PE ce EEES ENE J Se J m 500 4 5004 a A A 0 5 mg mL i 0 5 mg mL Haaa Ha Aa 1 2 3 4 5 6 7 8 9 min 0 1 2 3 4 5 6 7 8 9 min Column 4 6 mm x 150 mm 5 ym Mobile phase 50 0 1 TFA 50 ACN Flow 1 mL min Injection 10 pL Sample Doxepin Amitriptyline 0 5 50 mg mL in DMSO Figure 2 _Loadability comparisons for the basic compounds doxepin and amitriptyline using Agilent Prep C18 and a compe
4. 30 x 100 10 419910 302 419910 301 Prep 30 30 x 100 5 449905 302 449905 301 Prep 30 30 x 50 5 446905 302 446905 301 Prep 50 50 x 250 10 410910 502 410910 501 Prep 50 50 x 150 10 413910 502 413910 501 Prep 50 50 x 100 10 419910 502 419910 501 Prep 50 50 x 100 5 449905 502 449905 501 Prep 50 50 x 50 5 446905 502 446905 501 PrepHT Column Hardware Kit 820400 901 820400 901 PrepHT Guard Cartridges 2 pk 21 2 x 10 10 420212 902 420212 901 Prep HT Guard Hardware 820444 901 820444 901 Prep External Guard Hardware 420420 901 420420 901 Bulk packing 1kg 10 420910 902 420910 901 All PrepHT cartridge columns require hardware kit 820400 901 If a guard column is desired for the 21 2 mm ID columns the PrepHT Guard Hardware Kit 820444 901 is also required PrepHT Guard Hardware Kit 820444 901 If the guard column is used on a 30 mm or 50 mm ID column the External Guard column hardware kit 420420 901 is required Prep External Guard Hardware Kit 420420 901 For More Information For more information on our products and services visit our Web site at www agilent com chem Agilent shall not be liable for errors contained herein or for incidental or consequential damages in connection with the furnishing performance or use of this material Information descriptions and specifications in this publication are subject to change without notice Agilent Technologies Inc 2004 Printed in the USA July 9 2004 5989 1269EN
5. Agilent Prep LC Columns High Performance Preparative Columns for e e Maximum Sample Purification e o Technical Overview c00 e e e o e e e e e e Highlights under low pH 2 and high pH 10 pH conditions with basic compounds the most difficult sample High loadability for maximum sample type See Figure 1 purification Agilent Prep columns allow for a greater load with difficult basic compounds At pH 10 the best com petitive columns allowed for only 78 of the High Throughput 21 2 mm ID cartridges for fast sample load of the Agilent Prep C18 column before purification the peak width broadened substantially At pH 2 the Agilent Prep C18 column provided more than double the sample load for the same basic com e Ideal for use with the Agilent Purification System pounds While these comparisons were made using analytical 4 6 mm ID columns the results scale up to larger ID columns Agilent Prep columns provide superior loadability in any dimension e Easy scalability from 4 6 mm ID to 50 mm ID for rapid method development e Exceptional column stability from pH 2 10 Agilent Prep LC Columns A New Choice for Maximum Sample Purification High quality HPLC sample purification is critical in oan Agilent Prep C18 has higher loadibilityat pH 2 and 10 expanding drug discovery and drug development 100 pH 10 programs Agilent Prep LC columns available in mm pH2 both C18 and silica provide h
6. igh loadability for maximum sample purification with easy scalability to assure isolation and purification of milligrams to grams of desired drug compounds At the same time Agilent Prep LC columns provide the stabil ity high performance and flexibility to meet the most demanding purification needs 0 6 min os oO o 78 63 100 200 z ug Injection peak width S Agilent Prep C18 Competitor W C18 Competitor K C18 rior L ility over a Wi H upe 0 oadab ty over a de p Conditions pH 2 Conditions pH 10 Range Mobile phase 0 1 TFA in Water ACN 10 mM ammonia in Water ACN ratio was adjusted at ratio was adjusted at Sample loadability is determined by many factors PIE to hava samek OOO Bite to have sanek i ee Column 4 6 x 150 mm 5 pm including sample type acidic basic or neutral Flow 1 0 mL min resolution and mobile phase pH Therefore the Injection 20 pL best preparative columns provide excellent perfor Sample Papen MSO mance under a wide range of conditions The new Figure 1 Loadability comparison of the basic compound Agilent Prep columns provide superior loadability oxybutynin at pH 2 and 10 in DMSO using Agilent Prep C18 and two competitive columns one Agilent Technologies This superior loadability is achieved with high purity high surface area silica using a C18 bonded phase suitable for purification of a wide range of nonpolar and polar
7. ilent Prep columns pH or improving throughput and efficiency with a are also the best choice when using the Agilent 5 um particle size and short column length Agilent Purification System Now Agilent provides com Prep columns provide the flexibility for your plete solutions for the isolation and purification of demanding purification samples These reliable the widest range of compounds Table 3 Agilent Prep Columns Specifications Bonded phase Pore size Surface area Temp limits pH Range Endcapped Carbon load C18 100A 400 m g 60 C 2 10 Yes 24 Silica 100A 400 m2 g eds 1 8 N A N A N A Not Applicable Temperature limits are 60 C up to pH 8 40 C for pH 8 10 Temperature limits for bare silica are determined by the temperature of the mobile phase Table 4 Agilent Prep Columns Ordering Information Column description Size mm Particle size pm C18 Silica Scalar 4 6 x 250 5 440905 902 440905 901 Scalar 4 6 x 150 5 443905 902 443905 901 Scalar 4 6 x 100 5 449905 902 449905 901 Scalar 4 6 x 50 5 446905 902 446905 901 PrepHT cartridge columns require hardware kit 820400 901 PrepHT 21 2 x 250 10 410910 102 410910 101 PrepHT 21 2 x 150 10 413910 102 413910 101 PrepHT 21 2 x 150 5 443905 102 443905 101 PrepHT 21 2 x 100 5 449905 102 449905 101 PrepHT 21 2 x 50 5 446905 102 446905 101 Standard fittings no hardware required Prep 30 30 x 250 10 410910 302 410910 301 Prep 30 30 x 150 10 413910 302 413910 301 Prep 30
8. opment one can also immediately select a high performance preparative column based on estimated sample loads as shown in Table 2 Table 2 Typical Flow Rates and Column Loads Based on Column Size and Separation Separation type small molecules Normalized Easy Difficult Column ID flow rate gt 1 5 a gt 1 2 1 5 mm mL min mg mg 46 1 3 15 0 5 3 21 2 20 70 400 20 70 30 40 140 800 40 140 50 100 400 2000 100 400 Rapid High Throughput Purification High Throughput drug purification requires new column choices for preparative separations Shorter columns 50 100 mm long and smaller lt 10 micron particles are ideal Agilent Prep 21 2 mm ID columns are available with Agilent s Preparative Cartridge Hardware PrepHT This reliable cartridge hardware makes it easy to use short 50 and 100 mm long columns for rapid sample purification and then quickly change to a longer column length to conveniently increase sample load For even larger increases in sample load 30 mm and 50 mm ID columns with fixed end fittings can be used Cartridge columns 21 2 mm ID are also available in longer 150 and 250 mm lengths for more complex samples such as mix tures of natural compounds Longer 150 250 mm 30 and 50 mm ID columns are available for prod uct isolation Guard columns are easily integrated onto any of these columns providing superior pro tection of the prep column with complex samples Exceptional Column
9. titive column Easy Scalability for Rapid Method Development Column dimensions are available for preparative samples ranging from high throughput drug discovery samples to general lab scale prep sam ples Preparative columns are available in 21 2 30 and 50 mm ID with lengths ranging from 50 250 mm many with both 5 and 10 um particle sizes standard This wide variety of dimensions allows for optimizing the separation and maximiz ing the sample load on a scalar analytical 4 6 mm ID column before selecting the size of the prepara tive column needed Your separation and results will have the same high quality as you choose larger column dimensions for greater sample purification See the example shown in Figure 3 ID Flow rate Injection volume mm mL min nL 4 6 0 7 2 I T T T T T a a 0 2 4 6 8 10 12 min 42 4 A 21 2 14 87 JU T ry T T T eoo r 0 2 4 6 8 10 12 min 30 0 29 77 170 C N i T m T T T m e 7 0 2 4 6 8 10 12 min 50 8 85 37 488 pos L I T T T T T T T T T T T T 0 2 4 6 8 10 12 min Conditions System Agilent 1100 Purification System Detection UV 240 nm Columns 10 pm 150 mm Sample 1 Hydrocortisone Mobile phase 55 Water 45 ACN 2 Testosterone in MeOH at 1mg mL Temperature Room temperature Figure 3 An example of the excellent scalability of the Agilent Prep C18 column under nonoverload conditions making method transfer simple and predictable For more rapid method devel
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