Waters ACQUITY UPLC CSH Columns - Manual (Page 4)

[ Care and Use ManUal ]
ACQUITY UPLC CSH Columns 4
A listing of commonly used buffers and additives is given in Table 3. Additionally, the column lifetime will vary depending upon the operating
temperature, the type and concentration of buffer used. For example, the use of phosphate buffer at pH 8 in combination with elevated temperatures
will lead to shorter column lifetimes.
Table 2: Recommended pH and Temperature Limits for ACQUITY UPLC CSH Columns
Column Name
Particle Size
(µm)
Pore Diameter
(Å)
Surface Area
(m 2 )
pH Limits
Temperature Limits Ligand Density
(µmol/m 2 )
% Carbon
Low pH High pH
ACQUITY UPLC CSH C
18
1.7 135 185 1-11 80 45 2.3 15
ACQUITY UPLC CSH Phenyl-
Hexyl 1.7 135 185 1-11 80 45 2.3 14
ACQUITY UPLC CSH Fluoro-
Phenyl 1.7 135 185 1-8 60 45 2.3 10
Note: Working at the extremes of pH, temperature and/or pressure will result in shorter column lifetimes.
Table 3. Buffer Recommendations for Using ACQUITY UPLC CSH Columns up to pH 11
Additive/Buffer pKa Buffer Range Volatility
Used for
Mass Spec
Comments
TFA 0.3 Volatile Yes Ion pair additive, can suppress MS signal, used in the 0.02-0.1% range.
Acetic Acid 4.76 Volatile Yes Maximum buffering obtained when used with ammonium acetate salt. Used in 0.1-1.0% range.
Formic Acid 3.75 Volatile Yes Maximum buffering obtained when used with ammonium formate salt. Used in 0.1-1.0% range.
Acetate (NH
4
CH
2
COOH) 4.76 3.76 – 5.76 Volatile Yes Used in the 1-10 mM range. Note that sodium or potassium salts are not volatile.
Formate (NH
4
COOH) 3.75 2.75 – 4.75 Volatile Yes Used in the 1-10 mM range. Note that sodium or potassium salts are not volatile.
Phosphate 1 2.15 1.15 – 3.15 Non-volatile No Traditional low pH buffer, good UV transparency.
Phosphate 2 7.2 6.20 – 8.20 Non-volatile No Above pH 7, reduce temperature/concentration and use a guard column to maximize lifetime.
Phosphate 3 12.3 11.3 - 13.3 Non-volatile No Above pH 7, reduce temperature/concentration and use a guard column to maximize lifetime.
4-Methylmorpholine ~8.4 7.4 – 9.4 Volatile Yes Generally used at 10 mM or less.
Ammonia (NH
4
OH)
Ammonium Bicarbonate
9.2
10.3 (HCO
3
- )
9.2 (NH
4
+ )
8.2 – 10.2
8.2 – 11.3 Volatile
Volatile Yes
Yes Used in the 5-10 mM range (for MS work keep source >150 ˚C ). Adjust pH with ammonium
hydroxide or acetic acid. Good buffering capacity at pH 10.
Note: use ammonium bicarbonate (NH
4
HCO
3
), not ammonium carbonate ((NH
4
)
2
CO
3
)
Ammonium (Acetate) 9.2 8.2 – 10.2 Volatile Yes Used in the 1-10 mM range.
Ammonium (Formate) 9.2 8.2 – 10.2 Volatile Yes Used in the 1-10 mM range.
Borate 9.2 8.2 – 10.2 Non-volatile No Reduce temperature/concentration and use a guard column to maximize lifetime.
CAPSO 9.7 8.7 – 10.7 Non-volatile No Zwitterionic buffer, compatible with acetonitrile, used in the 1-10 mM range. Low odor.
Glycine 2.4, 9.8 8.8 – 10.8 Non-volatile No Zwitterionic buffer, can give longer lifetimes than borate buffer.
1-Methylpiperidine 10.2 9.3 – 11.3 Volatile Yes Used in the 1-10 mM range.
CAPS 10.4 9.5 – 11.5 Non-volatile No Zwitterionic buffer, compatible with acetonitrile, used in the 1-10 mM range. Low odor.
Triethylamine
(as acetate salt) 10.7 9.7 – 11.7 Volatile Yes Used in the 0.1-1.0% range. Volatile only when titrated with acetic acid (not hydrochloric or
phosphoric). Used as ion-pair for DNA analysis at pH 7-9.
Pyrrolidine 11.3 10.3 – 12.3 Volatile Yes Mild buffer, gives long lifetime.