UDC-Cholesterol™
Method Development Users Guide
CHOLESTEROL
Use this guide to predict your sample’s retention characteristics and
elution time on the Cogent UDC-Cholesterol Column (150 x 4.6mm ID).
Included are several examples of different classes of compounds of known
Octanol/Water Partition Coefficients (polarity). You can easily determine which
pH would be best to use for each of these compound classes with this column.
This User Guide will enable you to predict where your compounds should
elute for acids, bases and neutrals at three different pH’s.
How to Use This Guide
Step 1: Simply compare the Octanol/Water Partition Coefficient of your
acidic, basic or neutral compounds with the appropriate compounds shown.
Step 2: Read the pH condition at which you will have the best separation with the Generic, LCMS/compatible mobile phase for each
compound with this column.
Step 3: Predict the optimal pH for your mixture of compounds based
on the “retention maps” to resolve your complex mixtures of
acids, bases or neutral compounds
Octanol/Water Partition Coefficients (Log P) tend to be large for compounds with extended non-polar
structures (such as long chain or multi ring hydrocarbons) and small for compounds with highly polar
groups. Log P values provides a measure of hydrophilic vs. hydrophobic nature of your compound and
the UDC™ Cholesterol column.
MicroSolv Technology Corporation
101 Brighton Avenue, Long Branch, NJ 07740 USA
Ph. 1-732-229-3400 Fx. 1-732-229-2403
www.MicroSolvTech.com Email: [email protected]
—
—
—
Flow Rate: 3ml/min
Wavelength: 254 nm
100
90
CHOLESTEROL
% ACN
4-Hydroxy Benzoic
Acid (1)
80
Solvents
A: Buffer
B: 100% ACN
70
60
pH 2.18
50
40
—
—
—
30
20
10
0
1
2
3
Standards
4
5
6
7
8
9
10
11
8
9
10
11
TIME IN MINUTES
1 — 4-Hydroxyl Benzoic Acid
2 — Acetylsalicylic Acid
3 — 2-Hydroxy Benzoic Acid
100
90
80
% ACN
Acetylsalicylic
Acid (2)
70
60
50
pH 6.80
40
30
20
10
0
1
2
3
4
5
6
7
TIME IN MINUTES
2-Hydroxy
Benzoic Acid (3)
100
90
80
% ACN
—
—
—
RETENTION MAP
Acids
70
60
pH 8.00
50
40
30
20
10
0
1
2
3
4
5
6
7
8
9
10
11
TIME IN MINUTES
Octanol/Water Partition Coefficient
Uracil (1)
4-Hydroxy Benzoic Acid (2)
Acetylsalicylic Acid (3)
2-Hydroxy Benzoic Acidl (4)
-0.87
1.39
1.13
2.24
Octanol/Water Partition Coefficients (Log P) tend to be
large for compounds with extended non-polar structures
(such as long chain or multi ring hydrocarbons) and small
for compounds with highly polar groups. Log P values provides a measure of hydrophilic vs. hydrophobic nature of
your compound and the UDC™ Cholesterol column.
MicroSolv Technology Corporation
1-732-229-3400 • www.MicroSolvTech.com • [email protected]
90
% ACN
80
Nortriptyline (1)
70
pH 2.18
60
50
30
20
10
0
1
2
3
—
—
4
—
Flow Rate: 3ml/min
Wavelength: 254 nm
Solvents
A: Buffer
B: 100% ACN
40
5
6
7
8
9
10
11
8
9
10
11
TIME IN MINUTES
100
Standards
1 — Nortriptyline
2 — Imipramine
3 — Amitriptyline
90
Imipramine (2)
% ACN
80
70
60
50
40
pH 6.80
30
20
10
0
1
2
3
4
5
6
7
TIME IN MINUTES
100
90
Amitriptyline (3)
70
60
50
40
▼
Retention Time Reproducibility
When cycling pH 8.00 2.00 8.00
1
2
Original
8.3
9.4
First
8.2
9.3
Second
8.2
9.3
80
▼
—
—
—
100
CHOLESTEROL
% ACN
—
—
—
RETENTION MAP
Bases
3
9.9
9.8
9.8
pH 8.00
30
20
This requires 15 min equilibration each time.
10
0
1
2
3
4
5
6
7
8
9
10
11
TIME IN MINUTES
Octanol/Water Partition Coefficient
Nortriptyline (1)
Imipramine (2)
Amitriptyline (3)
1.63
3.49
2.18
Octanol/Water Partition Coefficients (Log P) tend to be
large for compounds with extended non-polar structures
(such as long chain or multi ring hydrocarbons) and small
for compounds with highly polar groups. Log P values provides a measure of hydrophilic vs. hydrophobic nature of
your compound and the UDC™ Cholesterol column.
MicroSolv Technology Corporation
1-732-229-3400 • www.MicroSolvTech.com • [email protected]
CHOLESTEROL
90
Dimethyl Phthalate (1)
% ACN
80
Solvents
A: Buffer
B: 100% ACN
70
pH 2.18
60
50
—
—
—
—
40
30
20
10
0
1
2
3
4
Standards
5
6
7
8
9
10
11
8
9
10
11
8
9
10
11
TIME IN MINUTES
1 — Dimethyl Phthalate
2 — Toluene
3 — Biphenyl
4 — Phenanthrene
100
90
Toluene (2)
80
% ACN
—
—
—
—
Flow Rate: 3ml/min
Wavelength: 254 nm
100
70
60
50
40
30
pH 6.80
20
10
Biphenyl (3)
0
1
2
3
4
5
6
7
TIME IN MINUTES
100
90
80
% ACN
—
—
—
—
RETENTION MAP
Neutrals & DMP*
Phenanthrene (4)
70
pH 8.00
60
50
40
30
20
10
0
1
2
3
4
5
6
7
TIME IN MINUTES
Octanol/Water Partition Coefficient
Octanol/Water Partition Coefficients (Log P) tend to be
Uracil (1)
-0.87
large for compounds with extended non-polar structures
Dimethyl Phthalate (2)
1.66
(such as long chain or multi ring hydrocarbons) and small
for compounds with highly polar groups. Log P values proToluene (3)
2.54
vides a measure of hydrophilic vs. hydrophobic nature of
your compound and the UDC™ Cholesterol column.
Biphenyl (4)
3.76
Phenanthrene (5)
4.35
* Added to mixture. Acts like a Neutral Compound.
MicroSolv Technology Corporation
1-732-229-3400 • www.MicroSolvTech.com • [email protected]