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Chromatographic hydrophobicity index (CHI) assay

Use CHI as a high throughput method for determining the lipophilicity of your compound.

Measurement of the chomatographic hydrophobicity index (CHI) is one of our Cloe Screen portfolio of in vitro ADME screening services. Cyprotex deliver consistent, high quality data with cost-efficiency that comes from a highly automated approach.

The chromatographic hydrophobicity index (CHI) as a measure of lipophilicity

Chromatographic hydrophobicity index (CHI) is a rapid gradient HPLC method for measuring lipophilicity and was first described by researchers from GlaxoSmithKline, UK¹.

The ADME characteristics of a compound are highly dependent on its lipophilicity.

Lipophilicity of a compound affects distribution into tissues, binding characteristics, absorption and elimination processes as well as solubility.

CHI correlates closely with traditional octanol-buffer distribution coefficient log D.

Rapid system which is cost effective and highly economical in terms of compound requirements.

Widely accepted method for determining lipophilicity.

‘The method is recommended as part of a protocol for high throughput physicochemical property profiling for rational drug design.’

¹Valko K, Bevan C and Reynolds DP. (1997) Anal Chem 69; 2022-2029.

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Protocol

CHI assay protocol

Analysis method	Reversed phase (C18) gradient with LC-MS/MS
Calibration	Literature accepted standards with known CHI values
Compound Requirements	20 μL of 10 mM solution in DMSO
Data Delivery	CHI_7.4

Lipophilicity is a critical determinant of the ADME properties of a drug - it can influence solubility, permeability, oral absorption, cell uptake, blood brain barrier penetration and metabolism.

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Data

Data from the Cloe Screen CHI assay

24 compounds were screened in Cloe Screen CHI_7.4 assay in quadruplicate on 4 separate occasions. These data are highly reproducible for a range of CHI values. Cloe Screen CHI_7.4 also correlates well with log D_7.4.

Compound name	Mean log D_7.4	Cloe Screen CHI_7.4
Phenylbutazone	0.55^(2,3)	57
Chlorpropamide	0.58^(2,3,4)	39
Trimethoprim	0.60⁽⁴⁾	39
Warfarin	0.76^{(2,3,4,5,6,7)}	46
Benzthiazide	1.12⁽⁴⁾	58
Chloroquine	1.28⁽⁴⁾	58
Carbamazepine	1.32^(3,4,8)	60
Dextromethorphan	1.40^(3,4)	60
Prednisone	1.48⁽³⁾	53
Midazolam	1.53⁽⁹⁾	76
Betamethasone	1.82⁽³⁾	60
Haloperidol	2.59^{(2,3,4,10,11)}	67
Ketoconazole	3.23^(3,4)	82
Nicardipine	3.77^(2,3,12)	102

Table 1

Cloe Screen CHI_7.4 data correlate closely with log D_7.4 data.

Figure 1

Cloe Screen CHI_7.4 data from 24
drugs show highly reproducible
intra- and inter-assay data. (error bars represent the standard deviation of 4 CHI_7.4 determinations).

Figure 2

Cloe Screen data show close correlation between CHI_7.4 and log D_7.4 for 14 known drugs (see table).

Q&A

Questions and answers on the chromatographic hydrophobicity index (CHI)

Please provide an overview of the Cloe Screen CHI assay.

CHI is an industry recognised high throughput method for determining lipophilicity. This method was originally developed by researchers at GlaxoWellcome¹³, and is based on reverse phase LC retention times. The method uses a literature accepted series of reference compounds with known CHI values¹⁴ to create a calibration line allowing CHI values for test compounds to be calculated according to their retention time.

Reference Compound	CHI
Theophylline	18.4
5-Phenyl-1H-tetrazole	23.6
Benzimidazole	34.3
Colchicine	43.9
8 Phenyl-theophylline	51.7
Acetophenone	64.1
Propiophenone	77.4
Butyrophenone	87.3
Valerophenone	96.4

Table 2

Table displaying the CHI values
for the reference compounds.

Figure 3

Graph showing the relationship between retention time and literature CHI values.

The gradient is calibrated by determining the retention time of each reference compound on a reverse phase LC system at pH 7.4. The test compounds are analysed on the same system. The equation generated from the calibration line for the reference compounds is then used to determine the unknown CHI value for the test compound.

What is the relationship between LogD and CHI?

CHI correlates closely with LogD_7.4 octanol/water partition coefficients as demonstrated in Figure 4.

Figure 4

The data below shows the relationship between LogD_7.4 and CHI for a set of 15 unrelated compounds.

How does lipophilicity influence the pharmacokinetic properties of a drug?

Lipophilicity is a key determinant of the pharmacokinetic behaviour of drugs. It can influence distribution into tissues, absorption and the binding characteristics of a drug, as well as being an important factor in determining the solubility of a compound.

How can CHI be used in predicting pharmacokinetics?

Lipophilicity significantly influences ADME properties and is widely used in drug discovery for QSAR (quantitative structure activity relationship) and QSPR (quantitative structure property relationship). This information is often used by our clients to rank compounds and determine which compounds to investigate in further studies.

What range of CHI values should I expect to see with my compound?

As CHI is a measure of retention of the test compound on the LC column a large range of values can be obtained. For compounds that are not retained on the column for any significant length of time a CHI value of <0 will be returned signifying very low lipophilicity. For compounds which are not easily washed off the column a CHI value of >120 is returned signifying very high lipophilicity. For any values in between the actual CHI value will be returned based on the calibration line of the reference compounds.

References

¹ Valko K et al. (1997) Anal Chem 69; 2022-9.
² Ritschel WA and Hammer GV. (1980) Int J Clin Pharmacol
Ther Toxicol 18 (7); 298-316.
³ Cyprotex internal Cloe Screen Log D Shake Flask data - for
protocol details see Cloe^® Screen Log D_7.4 Shake Flask
⁴ Cyprotex internal Cloe Select GLpKa data - for protocol
details see Cloe^®Select pK_a and log P
⁵ Murakami H et al. (2000) Am J Physiol Heart Circ Physiol
279 (3); H1022-8.
⁶ Camenisch G et al. (1998) Eur J Pharm Sci 6 (4); 317-24.

⁷ Yazdanian M et al. (1998) Pharm Res 15 (9); 1490-4.
⁸ Winiwarter S et al. (1998) J Med Chem 41 (25); 4939-49.
⁹ Greenblatt DJ et al. (1983) Br J Anaesth 55 (10); 985-9.
¹⁰ Nakashima E et al. (1987) J Pharm Sci 76 (1); 10-3.
¹¹ Yokogawa K et al. (1990) Pharm Res 7 (7); 691-6.
¹² Van de Waterbeemd H et al. (2001) J Comput Aided Mol
Des 15 (3); 273-86.
¹³ Valko K et al. (2001) Current Medicinal Chemistry 8, 1137-1146
¹⁴ Camurri G and Zaramella A (2001) Analytical Chemistry 73, 3716-3722

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