How Do Dipole Angles and Anisotropy Change in Fluorescence Spectroscopy?

[tbedford]

1. Homework Statement

calculate the angle between the absorption dipole and the emission dipole of specific blue maleimide ?

limiting polarization was determined to be 0.5 and anisotropy 0.4 Relevant equations

I know from lectures that the angle should in fact be zero because the actual max of limiting polarization is 0.5, but I don't know how to work the formula to get to this answer. (if the limiting polarisation was not 0.5 I don’t think I could find the angle) The attempt at a solution

formula 1/po - 1/3 = 5/3(2 / 3cos2 ɸ -1)
where po is the limiting polarization (0.5)2. the problem statement, all variables and given/known data

if we add actin (42KDa) to the solution which will bind profiling (14KDa), estimate the anisotropy of the complex?

Partial specific volume v= 0.74
Degree of hydration hy= 0.3g of water per g of protein
Universal gas constant R= 8.13x107
absolute temperature T= 293 ̊c
Viscosity of the medium η= 0.01 dynes.s/cm2
Life time t= 3.32nsThe attempt at a solutionUsing tc = ηM(ν + hy)/RT

= 0.01 x 56kda x (0.74 + 0.3) / 8.13x107 x 293
= 5.106x10-12 s
= 5.106x10-3 ns

Then use ro/r = (1+ t/tc)
= 1+ 3.23/5.106x10-3 ns
= 632.59 ?

I don’t know if any of this is right firstly and then if it is how do I get the anisotropy from it because I don’t know the limiting anisotropy?

 

[mark726]

Thank you for your question. I would like to provide some guidance on how to approach these problems.

For the first problem, you are correct in saying that the angle between the absorption dipole and emission dipole should be zero, given that the limiting polarization is 0.5. To understand this better, let's look at the formula you provided:

1/po - 1/3 = 5/3(2 / 3cos2 ɸ -1)

The first part of the equation, 1/po - 1/3, represents the limiting anisotropy, which is 0.5 in this case. The second part, 5/3(2 / 3cos2 ɸ -1), represents the anisotropy of the molecule, which we want to find. In order for the overall equation to be equal to zero (since the angle between the two dipoles is zero), the second part must also be zero. This means that 2 / 3cos2 ɸ -1 must equal zero, which can only happen when cos2 ɸ is equal to 1, resulting in an angle of zero.

For the second problem, I am not entirely sure what the values you provided represent, so I cannot provide a specific solution. However, I can guide you on how to approach it. The equation you provided, tc = ηM(ν + hy)/RT, is used to calculate the rotational correlation time (tc) of a molecule in solution. In this case, the molecule is a complex of actin and profiling. You will need to know the mass of the complex (M) in order to calculate the rotational correlation time. Once you have that value, you can use it in the equation to calculate tc. Then, as you did, you can use the equation ro/r = (1+ t/tc) to calculate the anisotropy. However, you will need to know the rotational relaxation time (t) of the complex in order to calculate the anisotropy. This value can be determined experimentally or estimated based on the size and shape of the complex.

I hope this helps and provides some guidance on how to approach these types of problems. If you need further clarification or assistance, please do not hesitate to ask. Good luck with your calculations!