DT034 Sensor Sheet

Sensor Description:

How it works:

Monochromatic light from an LED light source passes through a cuvette containing a solution sample, as shown in Figure 1. Some of the incoming light is absorbed by the solution. As a result, light of a lower intensity strikes a photodiode.

Transmittance and Absorbance

The amount of light that penetrates a solution is known as transmittance. Transmittance can be expressed as the ratio of the intensity of the transmitted light, It and the initial intensity of the light beam, Io, as expressed by the formula:

T=It/Io

The Colorimeter produces an output voltage, which varies in a linear way with transmittance, allowing a computer to monitor transmittance data for a solution. The transmittance of the sample varies logarithmically (base ten) with the product of three factors: E, the molar absorptivity of the solution, b, the cell or cuvette width, and C, the molar concentration.

log(l/T)= EbC

In addition, many experiments designed to use a colorimeter require a related measurement, absorbance. At first glance, the relationship between transmittance and absorbance would appear to be a simple inverse relationship. That is, as the amount of light transmitted by a solution increases, the amount of light absorbed might be expected to decrease proportionally. But the true relationship between these two variables is inverse and logarithmic (base 10). It can be expressed as:

A=log(l/T)

Combining the two previous equations, the following expression is obtained:

A = EbC

In effect, this formula implies that the light absorbed by a solution depends on the absorbing ability of the solute, the distance traveled by the light through the solution, and the concentration of the solution. For a given solution contained in a cuvette with a constant cell width, one can assume E and b to be constant. This leads to the equation:

A = kC (Beer's law)

Where k is a proportionality constant. This equation shows absorbance to be related directly to concentration and represents a mathematical statement of Beer's law. Beer's law is discussed in more detail later. In this guide, transmittance is expressed as percent transmittance or %T. Since T = %T/100, the formula can be rewritten as:

A = log(100/%T) or A = 2 - log%T

Wavelength Ranges:

You can select three LED light colors with the Colorimeter: Blue (470 nm or 4700 Angstram), green (565 nm or 5650 Angstram) and red (635 nm or 6350 Angstram). You can select one of these three nearly monochromatic colors using the wavelength selection knob on the top left of the colorimeter (see Figure 3). There are several ways you can decide which of the three wavelengths to use:

• Look at the color of the solution. Remember that the color of a solution is the color of light that passes through it. You probably want to use a different color of light that will be absorbed, rather than transmitted. For example, with a blue CuS04 solution, use the red LED (635 nm).
• Another quick method is to place a cuvette containing the solution in question in the Colorimeter and check to see which of the three wavelengths yields the highest absorbance (or lowest transmittance).
• Directions for most colorimetry experiments express a recommended wavelength. Use the closest of the three wavelengths on the colorimeter. Even if the LED wavelength is somewhat different, a Beer's law curve can usually be obtained at almost any wavelength in the vicinity of the recommended wavelength.

Calibration:

In order to properly work with the MultiLog, the Colorimeter should be defined as a "new sensor". Actually each of its colors should be defined as a "new sensor". Please refer to chapter 2.2.5 in the MultiLog User Manual, for instructions. We recommend that you recalibrate every time you perform a new colorimetry experiment or change the wavelength within an experiment. Though it is possible to save a calibration for future use, you will certainly see improved results if you recalibrate prior to doing a new experiment. A zero percent calibration is done with no light passing through a cuvette. The wavelength knob on the colorimeter is turned to "O% T" (see Figure 3). In this position the MultiLog can read data from the Colorimeter, but the light source is turned off. Since the light is off, it makes no difference if a cuvette is in the cuvette slot. A 100% calibration is done with the wavelength knob turned to select one of the three LED wavelengths. This turns on the red, green, or blue LED. A blank is placed in the cuvette slot. The blank is a cuvette containing the solvent used in the solution being studied, usually distilled water. The blank acts as a control by taking into account the small amount of light absorbed by the solvent and by the walls of the cuvette.

Follow these steps to calibrate the Colorimeter:

• First measure the Colorimeter output as a 0 to 5 Volt sensor.
• The wavelength knob is turned to the "0% T" position. The lid of the cuvette slot must be closed (with or without the "blank" cuvette in place). When the voltage value shown on the computer DB-Lab screen stabilizes, the user is asked to note this value. This value will later be defined as "O" (for 0% transmittance) .
• To do a 100% calibration, the blank is placed in the cuvette slot and the lid of the colorimeter is closed. The wavelength knob of the colorimeter is turned to one of the three LED colors. When the voltage value stabilizes, the user is asked to note a transmittance value. This value will later be defined as the "100" (for 100% transmittance).
• After this process is done proceed with the "Define Custom Sensor" dialog box (Para 2.2.5 in the MultiLog User's Manual). In the Volts section you should input the voltage values you noted and in the measured units you should enter "0" and "100" respectively.

In order for the DB-Lab software to be compatible with the right Colorimeter values, please choose from the DB-Lab software. During this process the MultiLog's LCD reads the actual voltage output from the Colorimeter, for both the 0% and 100%. These voltage values should be entered as the appropriate % values. This process should be done for each color.

Using Cuvettes with the Colorimeter

The Colorimeter is designed to use polystyrene cuvettes. Fifteen of these cuvettes and lids are supplied with the colorimeter. The cuvettes have a volume of approximately 4 ml. The cuvette Slot of the colorimeter is designed to give a snug fit to the cuvette and ensure that it is always in precisely the same position between the LED light source and photodiode. Two opposite sides of the cuvette are ribbed and are not intended to transmit the light from the LED. The two smooth surfaces are intended to transmit light. It is important to position the cuvette correctly in the colorimeter. We recommend this be done as shown in Figure 4, with the ribbed edges facing away from and toward you, and the smooth edges facing left and right. The light travels from left to right from the LED through the cuvette to the photodiode. It should also be noted that there is often a small variation in the amount of light absorbed by the cuvette if it is rotated 180? between trials. To avoid this, you should use a water-proof marker to make a reference mark on the right side of the top edge of the cuvette as shown in Figure 4. Or you can etch a reference mark using the hot tip of a soldering iron. Remind students to align this reference mark with the white reference mark on the top right side of the colorimeter each time they insert a cuvette.

Just like most spectrophotometer sample tubes, individual plastic cuvettes vary slightly in the amount of light they absorb. You may choose to ignore these differences. For most lab exercises, this variation will not have a noticeable effect on experimental results.
For best results, variation in light absorbed by individual cuvettes can be controlled either by using the same cuvette for all trials of a particular experiment or by matching a set of cuvettes.
The easiest and most reliable is the first method. If a student is going to use five trials for a Beer's law experiment, the five standard solutions can be transferred to the same cuvette for each trial. This requires that the cuvette be clean and dry after each trial or rinsed several times with the solution that will be added to it. This method takes very little time and successfully controls a potential variable. It also eliminates concerns over possible scratches that may eventually develop on a cuvette. The effect of the same small scratch is eliminated using the 100% calibration.
As an alternative, you may choose to match cuvettes. Matched cuvettes are a set of cuvettes that all absorb light (when empty) at approximately the same level. This involves more work on the part of the teacher, but saves time in student procedures. If students have 5 or 6 cuvettes with similar absorbance levels, then each sample can be added to a different cuvette, eliminating the drying or rinsing step described in the previous paragraph. To match a set of cuvettes, first calibrate the colorimeter using the method described in the section on calibration, Use a clean, dry cuvette for the 100% calibration instead of a distilled water blank. Put a reference mark on one of the clear sides of the cuvette so it is always oriented the same way in the cuvette slot. Place each cuvette in the batch in the Colorimeter and record transmittance values for each. When you are finished, group cuvettes according to similar %T values. Each of these groups represents a set of matched cuvettes.
Caps are supplied for the original 15 cuvettes. A cuvette may or may not have a cap on it when placed in the Colorimeter. The purpose of the cap is to prevent evaporation of solvent when an experiment is run over a period of several days. You may find it convenient to store standard solutions in capped cuvettes. If you purchase a replacement set of 100 cuvettes, 20 caps will be included. We felt teachers would probably not need to have one cap per cuvette. The caps can certainly be reused as cuvettes are replaced. It is very important that solutions be added to a cuvette to the proper depth. Our studies have shown that a "safe level" is between 2.2 and 3.5 mL of solution. When the cuvette is filled to the brim, its total volume is about 4.1 mL. Since the inside diameter of the cuvette is about 1.0 cm x 1.0 cm, this safe level can also be measured on the outside of the cuvette as 2.2-3.5 cm from the inside bottom of the cuvette. These levels are shown in Figure 5. As a reminder to students, these two levels could be marked with a water-proof marker on one of the ribbed sides of each cuvette.

What is it used for:

BEER'S LAW

• Crystal Violet:
  Dilute solutions of crystal violet yield a good Beer's law curve using the green LED (565 nm). A stock solution of 8 X 10^-5M crystal violet is prepared by adding 65.3 mg of solid crystal violet to enough water to yield 2 liters of solution. Dilute to obtain standard solutions.
• Copper Sulfate:
  Standard solutions that are 0.1, 0.2, 0.3 and 0.4M CuS04 will yield a good Beer's law curve at 635 nm (red LED). Or prepare a stock solution by adding 10 g of NH4NO3 to 10 mL of 0.1M CuSO4 and 90 mL of 0.20M NH3 (forms the Cu(NH3)4²⁺ complex ion) and dilute to obtain standard solutions.
• Food Coloring Solutions Red, Blue, Green:
  A less expensive alternative to using the solutions above is to prepare solutions using food coloring. You can obtain very good Beer's law curves using these solutions. Add about 6 drops of red, blue or green McCormick brand food coloring to 1 liter of water. The red solution can be analyzed using the blue LED (470 nm), the green solution with the blue LED (470 nm) or the red LED (635 nm), and the blue solution with the red LED (635 nm). Since the actual concentration of the solutions will not be known, refer to the original solution as "100%" and then dilute to 80, 60, 40, and 20%. Check the original solution to see that its absorbance is not greater than 0.550.

Specifications:

• Absorbance; 28% to 90%
• Transmittance : 28% to 90% ·
• Wavelengths:
  Blue: 470nm
  Green: 565nm
  Red: 635nm
• Cell width : 1cm
• Cell volume :3.5cc