BIST Ionic Modifier Preparation

BIST Mobile Phase Preparation
For use with BIST B and BIST B+ columns

The following procedures will produce a mobile phase (MP) solution containing 80% Acetonitrile (MeCN), 20% water, and 0.2% H2SO4. The resulting concentrations can be modified for your particular needs by adjusting the initial volumes of solution.

Chemicals:

HPLC-grade Acetonitrile (CAS #75-05-8)
HPLC-grade Water
HPLC-grade H2SO4 (CAS #7664-93-9)

Equipment:

1 L graduated cylinder
100 mL beaker,
1 L solvent bottle (qty. 2)
Analytical scale

The first step in making BIST-compatible Mobile Phases is to make 1 L of 0.2 % w/v H2SO4 in water and 1 L of 0.2% w/v H2SO4 in Acetonitrile. This will make further Mobile Phase preparations faster (and safer). The procedure below will be based on using 98% H2SO4 as the basis for the dilution. The numbers will change accordingly if you use 35% H2SO4

Preparation of 0.2% sulfuric acid (using 98% H2SO4)

  1. Ensure you are wearing proper PPE: gloves, lab coat, lab goggles and work under a fume hood.
  2. In the 1 L graduated cylinder, pour in about 500 mL of water.
  3. Weight  2.04 g of 98% H2SO4 reagent
  4. Add the H2SO4 into the 1 L graduated cylinder.
  5. Rinse the beaker a few times with about 50 mL of water adding washes to the cylinder
  6. Fill the graduated cylinder to the 1 L mark with water..
  7. Transfer the contents of the graduated cylinder into the 1 L solvent bottle  (This will be your acidic water stock solution.)
  8. In the 1 L graduated cylinder, pour in about 500 mL of acetonitrile.
  9. Weight  2.04 g of 98% H2SO4 reagent
  10. Add the H2SO4 into the 1 L graduated cylinder.
  11. Rinse the beaker a few times with about 50 mL of acetonitrile adding washes to the cylinder
  12. Fill the graduated cylinder to the 1 L mark with acetonitrile.
  13. Transfer the contents of the graduated cylinder into the 1 L solvent bottle (This will be your acidic acetonitrile stock solution.)
  14. Mix 800 mL of acidic acetonitrile and 200 mL of acidic water solution into the 1 L MP bottle to produce 80/20/0.2 acetonitrile/water/H2SO4 mobile phase.

 

Notes:
  • This procedure provides a slightly lower concentration of water and acetonitrile. The actual MP composition is close to 79.9/19.9/0.2, but for most practical purposes it can be written as 80/20/0.2.
  • We observed no significant changes in the acetonitrile sulfuric acid stock solution for a month of storage at room temperature. However there are some indications that acetonitrile can react very slowly with conc. sulfuric acid. So preferably the acetonitrile sulfuric acid stock solution needs to be keeped at lover temperature.

 

 

BIST Mobile Phase Preparation
For use with BIST A and BIST A+ columns

The following procedures will produce a mobile phase solution containing 80% MeCN, 20% water, and 5 mM N,N,N’,N’-Tetramethyl-1,3-diaminopropane (TMDAP) at pH 4.0. The resulting concentrations can be modified for your particular needs by adjusting the initial volumes of solution.

Chemicals:

HPLC-grade Acetonitrile (CAS #75-05-8)
HPLC-grade Water
99% N,N,N’,N’-Tetramethyl-1,3-diaminopropane (CAS #110-95-2)
HPLC-grade Formic acid (CAS #64-18-6)

Equipment:

500 mL graduated cylinder
500 mL beaker,
1 L solvent bottle (qty. 2)
Analytical scale
pH meter
Magnetic stirrer

100 mM TMDAP pH 4.0 buffer preparation:

  1. Ensure you are wearing proper PPE: gloves, lab coat, lab goggles and work under a fume hood.
  2. Weight 6.5 g of TMDAP into the beaker. 
  3. Add about 400 mL of water to the beaker
  4. Place the beaker on the stir plate with the stir bar in the beaker.
  5. Lower the pH probe into the beaker and turn on the stirrer.
  6. Slowly add formic acid into the beaker, monitoring the pH level until it reaches 4.0. It usually consumes about XX mL of formic acid in this step.
  7. Pour the contents of the beaker (except the stir bar) into the 500 mL graduated cylinder and fill the graduated cylinder to the 500 mL mark with water.
  8. Transfer the contents of the graduated cylinder into the 1L solvent bottle. (This will be your buffer stock solution)
  9. Mix in 1L bottle 150 mL of water, 50 mL of buffer stock solution and 800 mL of acetonitrile to produce 5 mM TMDAP/formate 80% acetonitrile MP.

 

Notes:
  • This MP can be prepared with a concentration of acetonitrile up to 90%. Higher concentration of acetonitrile makes MP non-homogeneous.