5.1.8: Solutions- instrumental analysis

CHEM 0103 Chemistry of the life sciences Lab #8 (follow-up to lab #7)

Where is that lab result? Concentration of an unknown

In the hospital, “the lab” usually means an analytical lab, and “lab result” means a determination of the concentration of molecules in a sample such as urine or blood. We will have four unknown solutions today (red, green, yellow, blue), and we will try to figure out whether they contain glucose or sodium chloride, and whether they are hypotonic, isotonic or hypertonic.

In order to analyze an unknown, we have to have standard solutions of known concentration to calibrate our assays. You will use the solutions you made 2 weeks ago. Today the entire lab will work together. Each group will be responsible for one out of four quantitative assays (experiments to figure out the concentration of a solute in a solution) and for one out of two qualitative assays. For the quantitative assay, you will create a standard curve relating concentration to measured quantity.

Then, each group will receive an unknown, and is responsible for figuring out whether it contains glucose or sodium chloride. Once we agree on the composition, each group will use their quantitative assay (and standard curve) to figure out its concentration.

Qualitative assays

Assay 1: Reaction with silver nitrate, AgNO3.

Safety: If \(\ce{AgNO3}\) solution comes into contact with your skin, you will see black stains appearing due to formation of elemental silver. Place tubes in a tube rack and use a pipette when transferring \(\ce{AgNO3}\) to avoid spills. If there is a spill, let the instructor clean it up or alert the instructor for help with cleaning up.

Procedure: Place 500 μL of the unknown (or standard) into a 1.5 mL centrifuge tube. Add a drop of \(\ce{AgNO3}\) and observe. Cap the tube, mix by inverting, and spin down in a centrifuge. Keep for comparison with other samples.

Interpretation: Chloride ions form an insoluble salt (AgCl) in the presence of Ag+ ions. If a precipitate forms, this is evidence for the presence of chloride ions (or other ions that form insoluble salts with Ag+).

Cleanup: Collect tubes containing \(\ce{Ag+}\) ions in the labeled hazardous waste container in the hood next to the \(\ce{AgNO3}\) reagent.

Assay 2: Benedict’s assay

Safety: As a heavy metal, copper and its compounds are toxic. Place tubes in a tube rack and use a pipette when transferring Benedict’s solution (a solution of \(\ce{CuSO4}\)) to avoid spills. If there is a spill, let the instructor clean it up.

Procedure: Place approx. 1 mL of Benedict’s solution into a 2 mL centrifuge tube, and add 100 uL of your sample. Place into the heat block for 5 min. and observe whether there is a color change. Centrifuge to collect the precipitate, if any.

Interpretation: \(\ce{Cu^2+}\) ions will be reduced when boiled in the presence of reducing sugars, forming a brown precipitate..

Cleanup: Collect tubes containing copper in the labeled hazardous waste container in the hood next to Benedict’s reagent.

Quantitative assays

Three groups will work on the glucose standard curves (density, refraction, and glucometer measurements). Three other groups will work on the NaCl standard curves (density, refraction, and conductivity measurements).

Density measurement

Place 20 mL of your sample into a clean 50 mL tube. Place on the balance, zero the display, and submerge a density probe (a marble on a fishing line) such that ist is completely submerged but does not touch the container. The mass corresponds to the mass of the displaced volume of solution. Calculate the density by dividing the measured mass by the known volume of the density probe. 

Index of refraction

Measure the refractive index of your unknown (or standard) in a refractometer. Open the sample compartment after moving the light source, clean the glass surface with a wipe, add 10 μL of your solution to the center of the glass surface, close the sample compartment and reposition the light source. You should see a sharp border between light and dark through the eyepiece. Turn the dial until the border is exactly on the cross hairs. Then, push the button on the side to reveal the scale. Use the bottom scale ("Brix scale") to read off your measurement.

Conductivity (NaCl solutions)

Place 3 mL of the unknown (or standard) into the 50 mL tube provided and add distilled water to 30 mL. Follow the instructions in this video. Briefly, insert the conductivity probe until it is submerged completely. Turn the drive button to read 4.0 and the sensitivity button counterclockwise to the limit. Turn the large (range) dial to maximize the darkened portion of the green display. Then, turn the sensitivity button clockwise to the limit, and use the drive button to maximize the darkened portion of the green display once more. Carefully read off the value, noting down both the number shown inside the instrument and the units and multiplier on the range dial. For example, a reading could be 7.6 x 100 uMhos. When you are done with the standard curve, test one glucose standard as a negative control.

Glucometer test (glucose solutions)

Dilute 20 μL of solution to 1 mL with water. Mix, and then use the commercial glucometer, inserting and dipping a new test strip as instructed. Some solutions might read "low"or "high", i.e. be outside of the range that is displayed. If this were blood samples of a diabetes patient, these low and high values either indicate faulty measurement or a medical crisis (abnormally low or high blood sugar levels). When you are done with the standard curve, test one NaCl standard as a negative control.