Waters PIC Reagents User Manual
[ Care and Use ManUal ]
[ Care and Use ManUal ]
PIC reagents
Contents
I. IntroduCtIon
II. PaIred-Ion ChromatograPhy
III. seleCtIng the reagent for your seParatIon
IV. usIng and storIng Waters PIC reagents
V. orderIng InformatIon
VI. Warranty/serVICe InformatIon
I. IntroduCtIon
Waters PIC® reagents allow simultaneous separation of acids, bases,
amphoteric, and neutral compounds on a reversed-phase column
by paired-ion chromatography (PIC). Separation of ionic species in
complex samples is easy with PIC reagents.
Use PIC A for separation of acids.
Use the PIC B series to separate bases.
Use Reagent D4 as a mobile phase modifier when separating basic
pharmaceutical compounds to improve peak shape and reduce reten-
tion times.
Low-UV PIC reagents are also available, for use in separations of
compounds that have chromophores below 240 nm.
II. PaIred-Ion ChromatograPhy
Paired-ion chromatography is a technique for performing ion-
exchange separations on a reversed-phase column. Hydrophobic
paired-ion reagents added to the mobile phase are adsorbed onto the
stationary phase. T here they enhance the interaction of oppositely
charged sample components with the stationary phase. A paired-ion
reagent:
• Increasestheretentionofoppositelychargedsolutes
• Decreasestheretentionofequallychargedsolutes
• Doesnotaffecttheretentionofunchargedsolutes
For a detailed description of the retention mechanism, see B.A.
Bidlingmeyer, S.N. Deming, W.P. Price, B. Sachok and M. Petrusek,
“Retention Mechanism for Reversed-Phase Ion-Pair Liquid Chromatog-
raphy”, Journal of Chromatography 186 (1979), 419-434.
III. seleCtIng the reagent for your seParatIon
Use PIC A Reagent to increase the retention of acids. Use the PIC B
Reagent to increase the retention of bases. Which of the four B
reagents to use depends upon your sample. Retention increases with
increasing reagent chain length. It is usually best to start with PIC B7.
If more retention is needed, use PIC B8. If less is needed, use PIC B5
or B6.
You can also use the concentration of organic modifier in your mobile
phase to manipulate the retention of all compounds, including neutral
compounds.
Table 1. PIC Reagents
Reagent Compound
PIC A Tetrabutylammonium phosphate
Low UV PIC A Tetrabutylammonium hydrogen sulfate
PIC B5 Pentane sulfonic acid
PIC B6 Hexane sulfonic acid
PIC B7 Heptane sulfonic acid
PIC B8 Octane sulfonic acid
Reagent D4 Dibutylammonium phosphate
PIC Reagents 1
[ Care and Use ManUal ]
[ Care and Use ManUal ]
Column: µBondapak C18, 3.9 x 300 mm steel column
Part Number: WAT027324
Mobile Phase: Methanol/Water 50:50 with PIC Reagent
Flow Rate: 2.0 ml./min
Detection: 254 nm
Sample: 1. Niacinamide
2. Pyridoxine
3. Thiamine
4. Riboflavin
PIC Separation of Water-Soluble B Vitamins
Compound or Class Reagent Compound or Class Reagent
Table 2. Reagents to Use for Common Compounds
Alkaloids Antitussives B
Cephaeline B Vitamins B5/B7
Emetinee B Calcium Leucovorin A
Ergot alkaloids B FD&C Dyes A
Analgesics/Antipyretics Folic Acid A
APC A Germicides
Aspirin A Benzalkonium Chloride B
Codeine B Cetylpyridinium Chloride B
Indomethacin A Imides A
Meperidine B Methotrexate A
Methadone B Petroleum Sulfonates A
Morphine B Surfactants
Propoxyphene B Quaternary amines B
Antibiotics Sulfonates A
Cephalosporins A or B
Penicillins A or B Amphetamine B
Antihistamines Ephedrine B
Chlorpheniramine B Epinephrine B
Diphenhydramine B Isoproterenol B
Methapyrilene B Naphazoline B
Phenindamine B Nylidrin B
Pyrilamine B Phenylephrine B
Tripelennamine B Phenylpropanolamine B
Pheniramine B
Sympathomimetics
(Decongestants)
Table 2 suggests the PIC reagent to use to separate the compo-
nents of some typical compounds on a C18 column with a water/
methanol eluent.
PIC Reagents 2
IV. usIng and storIng Waters PIC reagents
Waters PIC Reagents are premixed and buffered. Add 1 vial of reagent
to 1 L of mobile phase and filter.
Note: Crystals may form when vials are exposed to lower temperatures.
If crystals have formed, gently warm vial until crystals go back into
solution.
Store unopened reagent for up to one year at room temperature.
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