Use of Luminol in Forensic Analysis

Submitted by Patricia B. O'Hara on Monday, 11/16/2009, at 2:12 PM

Day 6:  Light and Molecules...Use of Luminol in Forensic Analysis

O’Hara, P.; Engelson, C.; St. Peter, W. J. Chem. Educ. 2005,82, 49

OBJECTIVE: To detect trace amounts of an oxidant such as hemoglobin using chemiluminescence and thereby detect which common lab instrument was used as a murder weapon. 

 BACKGROUND: Chemiluminescence is the emission of light as a result of a chemical reaction.  Ancient documents almost two thousand years old record Pliny the Elder’s observation that a walking stick rubbed with the luminescent jellyfish, pulmo marinus, would “light the way like a torch”.  The term chemiluminescence was first used in 1888 by Eilhardt Weidemann in his studies of reactions that produced light without heat or “cold light”.  We have come to learn that this light is derived from unstable high energy intermediates in some chemical reactions.

A + B high energy intermediate  → products + light                       

In this experiment, you will use a basic solution of luminol, 3-amino­phthal­hydrazide, to react with an oxidant (bleach) to generate the cold blue light characteristic of this reaction.  The molecular structure of luminol and proposed reaction are shown in the attached file.   The cold blue emission of luminol is best viewed in a darkened room. Chemiluminescence persists for seconds to minutes depending on the concentration of oxidant, and several other factors. 

FORENSICS:  Crime scene investigators, police detective specialists who have advanced training in forensic science, consider luminol to be an important tool in their investigation of crime scenes.   Blood contains an oxidant, hemoglobin, powerful enough to oxidize luminol for extended periods of time after a crime has been committed and the reaction is sensitive enough to detect the presence of blood even after the article contaminated with blood has been washed.  A number of prime time television shows have made use of this chemiluminescent reaction as part of their crime scene investigations.  A quick search of newspapers like the New York Times reveals 38 references to the use of luminol in the last two years.   It is important to emphasize here that a positive luminol result is suggestive, but not proof that blood is present on the sample.  Should detectives wish to make a conviction, further evidence is necessary, since there are many other molecules besides blood (such as bleach, or any other oxidizing material) that can react with luminol.  It is also a destructive assay.  Blood assayed by luminol cannot also be used for DNA analysis, so research is active in the forensics field to develop an indicator for blood that is as sensitive as luminol but more specific and non-destructive.

THE SCENE:  A veritable cantankerous professor of chemistry has been murdered in her lab.  There are multiple stab wounds on the body, but it appears as though the murder weapon has vanished.  Several other pieces of laboratory equipment (spatula, pipettor, stirring rod and a thermometer) are scattered about on the floor.  Could one of them be the murder weapon?

OBJECTIVE:   It is your job to use the chemiluminescence of luminol to discover which pieces of laboratory equipment may have been exposed to “blood”. (Note: no real blood has been used in this experiment.)

Materials List and Warnings, see attached file.

Procedure:

1.   Prepare a basic solution of luminol by dissolving 5.0 g of sodium carbonate in 100 mL of distilled water.   Add 0.1 g of luminol reagent, and stir or swirl to dissolve. Transfer contents to a spray bottle.

2.   Sodium perborate is not indefinitely stable.  Swirl 0.7 g of sodium perborate catalyst to dissolve in the the 100 mL of luminol solution in your spray bottle just before analysis.   

3.   Darken the room and spray the luminol solution onto the potentially contaminated pieces of laboratory equipment to discover which produce the cold blue emission of luminol.  You may need a few sprays to see the luminescence but you will most certainly have material left over.

 

Record your observations below: 

Reaction with spatula

Reaction with pipette

Reaction with stir rod. 

Reaction with thermometer

 

 

 

 

 

 

 

 

 

 

ANALYSIS:

  1. Which pieces of equipment are contaminated?

 

  1. Review the relationships of color to wavelength.  What is the approximate wavelength of light emitted by the luminol reaction? 

 

  1. Using the relationships below, calculate the change in energy (per photon and per mole) as luminol intermediates return to the ground state by emitting a photon.

 

E = h n per photon                                           c =  l  n

            E = energy                                                                   c (speed of light) = 2.998 x 108 m/s

            h (Planck’s constant)  = 6.626 x 10-34 J.s                     l = wavelength

            N (Avogadros number) =  6.02 x 1023 mole-1                  n  = frequency               


ANSWER SHEET 

 

For example:

Record your observations below: 

Reaction with spatula

Reaction with pipette

Reaction with stir rod. 

Reaction with thermometer

Light blue emission lasts several sec (not indefinitely)

No reaction after repeated spraying

 

No reaction after repeated spraying

 

No reaction after repeated spraying

 

 

ANALYSIS:

  1. Which pieces of equipment are contaminated?  On the basis of these results, it appears as though the spatula was the only piece of equipment to be exposed to “blood”

 

  1. Review the relationships of color to wavelength.  What is the approximate wavelength of light emitted by the luminol reaction?  Blue light is approximately 400 nm = 4.00 x 10-7m

 

  1. Using the relationships below, calculate the change in energy (per photon and per mole) as luminol intermediates return to the ground state by emitting a photon. 

E = h n per photon                                          c =  l  n

            E = energy                                                                   c (speed of light) = 2.998 x 108 m/s

            H (Planck’s constant)  = 6.626 x 10-34 J.s                    l = wavelength

            N(avogadros number) =  6.02 x 1023mole-1                        n  = frequency             

 

Per photon: E = (6.626 x 10-34 Js)(2.998 x 108 m/s) / (4.00 x 10-7m)  =      4.97 x 10-19 J 

Per mole:    E= 4.97 x 10-19 J x 6.02 x 1023 mole-1 = 2.99 x 105 J/mole          =   299 kJ

Forensics Lab

Submitted by Patricia B. O'Hara on Monday, 11/16/2009, at 2:14 PM