Paper Chromatography Lab

Introduction

Chromatography is a method of separating mixtures based on the properties of the substances being separated. Separation by chromatography generally requires two phases of matter - a stationary phase and a mobile phase. The stationary phase is usually a solid, in this case the paper, which allows some substances to pass through easily and holds others back. The mobile phase is either a gas or liquid (the water in this lab) that moves through the stationary phase, ‘carrying’ the substances to be separated.

Chromatography was invented in the early 1900s by Michael Tswett to separate plant pigments. Today many variations of chromatography are used to separate mixtures of substances so that they can be identified. The food industry uses gas chromatography to separate and identify the compounds that give foods their aromas. Some of the compounds discovered are then synthetically made in the laboratory and used for artificial flavors and fragrances. Gas-liquid chromatography is used for drug testing and for determining the percent of alcohol in a blood sample. One variation of chromatography that is becoming increasingly important is electrophoresis used for DNA fingerprinting. Forensic scientists use DNA fingerprinting to help identify people and solve crimes.

In this laboratory experiment you will be using paper chromatography to separate several mixtures.

Procedure (Holt text p.848-849)

1.  Obtain a piece of filter paper about 12cm in diameter. Use a graphite pencil to sketch a circle about the size of a quarter in the center of the filter paper.

2.  Use a graphite pencil to write a number (#1-6) around the inside of this circle for each sample to be tested. The six unknown mixtures will be ink.

3.  Label the unknown markers #1-6. Use each marker and draw a short line (~0.3cm) on the pencil circle at each marker’s corresponding number (#1-6).

4.  Use the pencil to poke a small hole in the center of the spotted and labeled filter paper. Insert a wick through the hole. A wick can be made by rolling a small piece of filter paper into a cylinder.

5.  Fill a Petri dish two thirds full of solvent (water in this case).

6.  Set the bottom of the wick in the solvent so that the filter paper rests on the top of the Petri dish.

7.  When the solvent is 2cm from the outside edge of the paper, remove the paper from the Petri dish and allow the chromatogram to dry.

Data

A. Create a diagram that can be used to record and easily compare your qualitative observations for each of the substances that are being separated. A color diagram of your experiment with written descriptions will suffice. Be neat and label your diagram, being detailed in your descriptions.

B. Re-draw and extend the sample data table below to record several quantitative observations for each of the substances. (Reminder: all data tables should be drawn neatly using a ruler and must include a complete, descriptive title, and subtitles for columns and rows.) Include space to record observations for an unknown marker.

C. Calculate the retention factor (Rf) values for each of the substances and record.

Rf = distance substance moved

distance mobile phase moved

Calculation of Retention Factor Values / Additional Quantitative Observations
Marker used / Color / Distance Substance Moved (in mm.) / Distance Mobile Phase Moved (in mm.) / Rf Value
1) / a)
b)
c)
(etc)
2) / a)
b)
c)
(etc)
3) (etc)

Data Table: ______

Questions (write answers in lab report)

1)  How many different substances were found in each of the different pens? Describe each pen in detail.

2)  The number/letter of your unknown is ______. What is its most likely identity?

3)  What is chromatography?

4)  Name at least 3 properties that can be used to separate substances by chromatography.

5)  Define retention factor. Explain its usefulness.

6)  Suggest some possible reasons explaining three qualitative observations you made (e.g. what reason(s) could explain the lack of color separation for one of the markers? etc...).
i) Observation:
Explanation:
ii) Observation:
Explanation:
iii) Observation:
Explanation:

7)  Some components of ink are minimally attracted to the stationary phase and very soluble in the solvent (mobile phase). Where are these components most likely to be located on the filter paper during chromatography?

8)  Predict the results of forgetting to remove the chromatogram from the water in the petri dish until the next day.

9)  How could this method be used to help identify an unlabeled marking pen?