Mariotte flask

A constant hydrostatic pressure during chromatography is achieved if a Mariotte flask (Fig. 3.3) is used. Regulate the flow rate by varying Ah and/or by a pinch clamp at the outlet. 

Mariotte flask A reservoir that maintains a constant hydrostatic pressure for liquid chromatography. 

Fig[ure 5-7. Operating or hydrostatic pressure for various column assemblies. A and B pressure is measured between the free eluent surface in the column or the reservoir and the end of the outlet tubing. C and D pressure is measured from the bottom of the air inlet tube in the Mariotte flask to the end of the outlet tubing, no matter whether the flow is downward (C) or upward (D). 

Fignre 5-8. Schematic representation of a Mariotte flask in operation. 

Assemble on the fraction collector a Mariotte flask containing 400 ml buffer A and adjusted to yield a hydrostatic head of 58 cm. 

When this solution reaches the gel surface very gently fill the column with buffer A, replace the column cap, and connect the Mariotte flask to the column. Under these conditions the column will exhibit a flow rate of approximately 5 ml/1.5 minutes. This is much faster than would usually be used and results in considerable dilution of the sample. However, it demonstrates well the technique s potential in a short time. Note also that the two materials completely separate in about 3 to 4 cm of gel. Therefore, a column of only 3 to 5 cm would be needed for this particular sample size and composition. Separations of this type provide a good illustration of the column capacities needed for desalting applications such as those discussed in Chapter 10. Under these conditions a 0.5 ml protein sample could be desalted in less than 10 minutes. 

In many cases flow under gravity is satisfactory and in this case a pump is not needed. A constant head may be obtained using a Mariotte flask (e.g. Fischer, this series, 1969) such as that obtainable from Pharmacia Fine Chemicals AB. The principle is shown in Fig. 4.5. 

Fig. 4.5. Mariotte flask. A constant pressure head device.

An eluent container can be a Mariotte flask that produces constant hydrostatic pressure, and thus it can serve for the generation of the gravitational eluent transport in the chromatographs of the first generation. Simple solvent tanks, or even open vessels, are used as eluent containers in the instruments provided with efficient pumping systems. The preferential evaporation effects, however, should be prevented when working with multicomponent, mixed eluents. 

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