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Clarke's Analysis of Drugs and Poisons

Basic nitrogenous drugs

A. H. Stead et al., Analyst, 1982, 107, 11061168 and R. A. de Zeeuw et al., Thin-layer
Chromatographic Rf Values of Toxicologically Relevant Substances on Standardized Systems:
Report XVII of the DFG Commission for Clinical-Toxicological Analysis, 2nd Edn, VCH,
Weinheim, 1992.

System TA

Plates: Silica gel G, 250 m thick, dipped in, or sprayed with, 0.1 M potassium
hydroxide in methanol, and dried.
Mobile phase: Methanol:strong ammonia solution (100:1.5).
Reference compounds: Atropine Rf 18, Codeine Rf 33, Chlorprothixene Rf 56,
Diazepam Rf 75.

System TB

Plates: Silica gel G, 250 m thick, dipped in, or sprayed with, 0.1 M potassium
hydroxide in methanol, and dried.
Mobile phase: Cyclohexane:toluene:diethylamine (75:15:10).
Reference compounds: Codeine Rf 06, Desipramine Rf 20, Prazepam Rf 36,
Trimipramine Rf 62.

System TC

Plates: Silica gel G, 250 m thick, dipped in, or sprayed with, 0.1 M potassium
hydroxide in methanol, and dried.
Mobile phase: Chloroform:methanol (90:10).
Reference compounds: Desipramine Rf 11, Physostigmine Rf 36, Trimipramine
Rf 54, Lidocaine Rf 71.

System TL

Plates: Silica gel G, 250 m thick, dipped in, or sprayed with, 0.1 M potassium
hydroxide in methanol, and dried.
Mobile phase: Acetone.
Reference compounds: Amitriptyline Rf 15, Procaine Rf 30, Papaverine Rf 47,
Cinnarizine Rf 65.

System TAE

Plates: Silica gel G, 250 m thick.


Mobile phase: Methanol.
Reference compounds: Codeine Rf 20, Trimipramine Rf 36, Hydroxyzine Rf 56,
Diazepam Rf 82.

System TAF

Plates: Silica gel G, 250 m thick.


Mobile phase: Methanol:n-butanol (60:40) and 0.1 mol/L NaBr.
Reference compounds: Codeine Rf 22, Diphenhydramine Rf 48, Quinine Rf 65,
Diazepam Rf 85.

Location reagents for systems TA, TB and TC

Ninhydrin spray
Spray the plate with the reagent and then heat in an oven at 100 for 5 min. Violet or pink spots
are given by primary amines and yellow colours by secondary amines.

FPN reagent
Red or brown-red spots are given by phenothiazines and blue spots by dibenzazepines. This
reagent may be used to overspray a plate which has been previously sprayed with ninhydrin
spray.

Dragendorff spray
Yellow, orange, red-orange, or brown-orange spots are given by tertiary alkaloids. This reagent
may be used to overspray a plate which has been previously sprayed with ninhydrin spray and
FPN spray.

Acidified iodoplatinate solution


Violet, blue-violet, grey-violet, or brown-violet spots on a pink background are given by tertiary
amines and quaternary ammonium compounds. Primary and secondary amines give dirtier
colours. This solution may be used to overspray a plate which has previously been sprayed with
ninhydrin spray, FPN reagent and Dragendorff spray.

Mandelins reagent
This reagent is preferably poured onto the plate because of the danger of spraying concentrated
acid. Many different colours are given with a variety of drugs (see Chapter 19 and the Index of
Colour Tests).

Marquis reagent
This reagent is preferably poured onto the plate because of the danger of spraying concentrated
acid. Black or violet spots are given by alkaloids related to morphine. Many different colours are
given with a variety of drugs (see Chapter 19).

Acidified potassium permanganate solution


Yellow-brown spots on a violet background are given by drugs with unsaturated aliphatic bonds.

Rf values
Rf values for drugs in these systems will be found in drug monographs and in the Indexes of
Analytical Data in Volume 2; they are also included in the systems for specific groups of drugs
which follow.

Acidic and neutral drugs

A. H. Stead et al., Analyst, 1982, 107, 11061168 and R. A. de Zeeuw et al., Thin-layer
Chromatographic Rf Values of Toxicologically Relevant Substances on Standardized Systems:
Report XVII of the DFG Commission for Clinical-Toxicological Analysis, 2nd Edn, VCH,
Weinheim, 1992.

System TD

Plates: Silica gel G, 250 m thick.


Mobile phase: Chloroform:acetone (80:20).
Reference compounds: Paracetamol Rf 15, Clonazepam Rf 35, Secobarbital Rf
55, Methylphenobarbital Rf 70.

System TE

Plates: Silica gel G, 250 m thick.


Mobile phase: Ethyl acetate:methanol:strong ammonia solution (85:10:5).
Reference compounds: Sulfadimidine Rf 13, Hydrochlorothiazide Rf 34,
Temazepam Rf 63, Prazepam Rf 81.

System TF
Plates: Silica gel G, 250 m thick.
Mobile phase: Ethyl acetate.
Reference compounds: Sulfathiazole Rf 20, Phenacetin Rf 38, Salicylamide Rf
55, Secobarbital Rf 68.

System TAD

Plates: Silica gel G, 250 m thick.


Mobile phase: Chloroform:methanol (90:10).
Reference compounds: Hydrochlorothiazide Rf 11, Sulfafurazole Rf 33,
Phenacetin Rf 52, Prazepam Rf 72.

Location reagents for systems TD, TE and TF

Acidic drugs

Van Urk reagent


Spray the plate with the reagent and then heat in an oven at 100 for 5 min. Yellow spots are
given by sulfonamides and by meprobamate, blue spots are given by ergot alkaloids, and pink or
violet spots are given by some other compounds, e.g. phenazone.

Ferric chloride solution


Blue or violet spots are given by phenols. This solution may be used to overspray a plate which
has been previously sprayed with Van Urk reagent.

Mercurous nitrate spray


Barbiturates give dark spots which fade slowly; with some dilute solutions the spots fade rapidly.

Acidified potassium permanganate solution


Yellow-brown spots on a violet background are given by drugs with unsaturated aliphatic bonds,
e.g. secobarbital. This solution may be used to overspray a plate which has been previously
sprayed with mercurous nitrate spray.

Neutral drugs

Furfuraldehyde reagent
Violet to blue-black spots are given by some neutral compounds, e.g. carbamates.

Acidified iodoplatinate solution


This solution may be used to overspray a plate which has been previously sprayed with
furfuraldehyde reagent.

Rf values
Rf values for drugs in these systems will be found in drug monographs and in the Indexes of
Analytical Data in Volume 2; they are also included in the systems for specific groups of drugs
which follow.

Note
It is worth noting that system TE can be used for acidic, neutral and basic drugs. Furthermore,
systems TC and TAD use the same mobile phase so that acidic, neutral and basic drugs can be run
in the same tank, although on separate plates. It should also be noted that the above systems for
basic nitrogenous drugs are also able to separate neutral drugs if the latter are present in the
sample or in the basic extract thereof.

Finally, the Index of Colour Tests lists colour reactions with TLC spray reagents for
approximately 250 compounds and may therefore serve as an indication of colour reactions
specific to certain classes of compounds.
General screening systems

The TLC systems listed below (Systems TAJ, TAK and TAL) were developed primarily by
Professor George Maylin, New York State Racing Wagering Board, Drug Testing Programme, as
well as System TAM listed under steroids.

System TAJ

Plates: Silica gel G, 250 m thick.


Mobile phase: Chloroform:ethanol (90:10).

System TAK

Plates: Silica gel G, 250 m thick.


Mobile phase: Chloroform:cyclohexane:acetic acid (4:4:2).

System TAL

Plates: Silica gel G, 250 m thick.


Mobile phase: Chloroform:methanol:propionic acid (72:18:10).

Location reagents for systems TAJ, TAK, TAL and TAM

Cupric chloride

Dragendorff spray

Fearon's reagent

Ferric chloride, ethanol and sulfuric acid

Fluorescamine

Gibb's reagent

Conc. hydrochloric and ethanol

Iodine

Mandelin's reagent

Modified Ehrlich's reagent

Sodium nitrite

Ninhydrin