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Beyond Beer's Law: Why the Index of Refraction Depends (Almost) Linearly on
Concentration

Article  in  ChemPhysChem · February 2020

DOI: 10.1002/cphc.202000018

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 DOI: 10.1002/cphc.202000018 Communications

1
2
3 Beyond Beer’s Law: Why the Index of Refraction Depends
4
5
(Almost) Linearly on Concentration
6
7
Thomas G. Mayerhöfer,*[a, b] Alicja Dabrowska,[c] Andreas Schwaighofer,[c] Bernhard Lendl,[c]
8 and Jürgen Popp[a, b]
9
10
Beer’s empiric law states that absorbance is linearly propor- as it was not discussed nor provided in this form in Kayser’s
11
tional to the concentration. Based on electromagnetic theory, handbook of spectroscopy,[3] which was the reference work at
12
an approximately linear dependence can only be confirmed for this time. When Max Planck derived his particular kind of
13
comparably weak oscillators. For stronger oscillators the dispersion theory and employed it to compare the results with
14
proportionality constant, the molar attenuation coefficient, is Beer’s law in 1903,[4] he was most probably unaware of
15
modulated by the inverse index of refraction, which is itself a Equation (1), otherwise he probably would have derived it from
16
function of concentration. For comparably weak oscillators, the dispersion theory. Recently, this derivation was carried out, first
17
index of refraction function depends, like absorbance, linearly from dispersion theory[5] and, subsequently, from simple
18
on concentration. For stronger oscillators, this linearity is lost, electromagnetic theory.[6] One of the main results of the
19
except at wavenumbers considerably lower than the oscillator derivation is that the molar attenuation coefficient is inversely
20
position. In these transparency regions, linearity between the proportional to the index of refraction, which is itself a function
21
change of the index of refraction and concentration is of concentration. Concerning the nature of this dependence, as
22
preserved to a high degree. This can be shown with help of the was already stated in Ref. [5], a law comparable to Beer’s law,
23
Kramers–Kronig relations which connect the integrated absorb- but for the index of refraction function, can be derived. In ref.[5]
24
ance to the index of refraction change at lower wavenumbers the nature of this dependence was not investigated in detail. As
25
than the corresponding band. This finding builds the founda- we will show in the following, the use of the index of refraction
26
tion not only for refractive index sensing, but also for new variation instead of the absorbance to investigate concentration
27
interferometric approaches in IR spectroscopy, which allow may have several advantages.
28
measuring the complex index of refraction function. To derive the concentration dependence of the complex
29
index of refraction, it is possible to start from simple electro-
30
magnetic theory. Accordingly, the macroscopic polarization ~ P is
31
related to the dipole moment ~ p induced in an atom or a
32
The exact origin of the form of Beer’s law, as we employ it molecule by the following relation:
33
nowadays[1,2]
34 !
35
P ¼ N �~
p (2)
Að~vÞ ¼ e* ð~vÞ � c � d (1)
36
Here, N is the number of dipole moments per unit volume
37
wherein e* ð~vÞ is the molar attenuation coefficient, c the (which is equal to the molar concentration multiplied by
38
concentration and d the sample thickness, remains unclear. It Avogadro’s constant, N = NA·c). Inherent to Equation (2) is the
39
seems that it was not employed in this form before about 1900, assumption that there are no interactions between the micro-
40
scopic dipoles, i. e. there are no chemical interactions. On the
41
[a] Dr. T. G. Mayerhöfer, Prof. Dr. J. Popp
other hand, macroscopically, for scalar media, the polarization is
42
Spectroscopy/Imaging related to an applied electric field ~
E [Eq. (3)]:
43 Leibniz Institute of Photonic Technology
44 Albert-Einstein-Str. 9, 07745 Jena, Germany ~
P ¼ e0 ðer 1Þ~
E ¼ e0 c~
E (3)
45 [b] Dr. T. G. Mayerhöfer, Prof. Dr. J. Popp
Institute of Physical Chemistry and Abbe Center of Photonics
46 Friedrich Schiller University wherein er is the relative dielectric function, c the electric
47 Helmholtzweg 4, 07743 Jena, Germany
E-mail: [email protected]
susceptibility and e0 the vacuum permittivity. The use of a scalar
48
[c] A. Dabrowska, Dr. A. Schwaighofer, Prof. Dr. B. Lendl dielectric function implies that the polarization is always co-
49 Institute of Chemical Technologies and Analytics oriented to the electric field and that the material is isotropic
50 Technische Universität Wien
Getreidemarkt Wien, 9/164, 1060 Vienna, Austria
and homogenous in relation to the resolution limit of light.
51
Supporting information for this article is available on the WWW under Using the relation between electric susceptibility and polar-
52
https://doi.org/10.1002/cphc.202000018 izability a, c ¼ N � a=e0 , and assuming additionally that there
53
© 2020 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This are no local field effects, we find that [Eq. (4)]:
54 is an open access article under the terms of the Creative Commons Attri-
55 bution Non-Commercial NoDerivs License, which permits use and distribu-
56 tion in any medium, provided the original work is properly cited, the use is
non-commercial and no modifications or adaptations are made.
57

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 Communications

1 NA � a
c¼c (4)
2 e0
3
4
A similar equation relates in Raman spectroscopy the Raman
5
tensor and the change of polarizability of a molecule or a unit
6
cell during a vibration. In contrast to Raman spectroscopy,
7
however, in UV-Vis and IR-spectroscopy we have to invoke
8
Maxwell’s wave equations to understand the changes of the
9
intensity due to absorption. Assuming that our medium is not
10
only scalar but perfectly homogenous (in particular nearfield
11
interactions/electromagnetic coupling are excluded[7]), then the
12
simple and well-known relation er ¼ n ^2 follows,[8] wherein n
^ is
13
the complex index of refraction. Accordingly, it follows:
14
15 rffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
pffiffiffiffiffiffiffiffiffiffiffi N �a
16 ^ ¼ n þ ik ¼
n 1þc¼ 1þc A (5)
e0
17
18
Focusing on the (real) index of refraction n and assuming
19 pffiffiffiffiffiffiffiffiffiffiffi
that the electric susceptibility c � 1, so that 1 þ c � 1 þ c=2,
20
we find: Figure 1. Upper panel: Beer’s law (black line) compared with concentration
21 dependence of the normalized integrated absorbance (from 1600–
22 1800 cm 1, red line), the normalized change of the index of refraction at
NA � a0 ð~vÞ 1500 cm 1 (green line), the normalized change of the index of refraction at
23 nð~vÞ ¼ 1 þ c (6)
2e0 1693 cm 1 (blue line), and the absorbance at the oscillator position
24 (1700 cm 1, pink line). Center panel: Index of refraction function. Lower
25 panel: Index of absorption function for different concentrations of 0.5, 5, 10,
Here, a0 ð~vÞ is the real and wavenumber dependent part of 25 and 50 mol L 1 (S*2 = 4900 L/(mol cm2), ~v0 ¼1700 cm 1 γ = 20 cm 1).
26
the polarizability. Therefore, as long as the electric susceptibility
27
is small, a linear relation between the change of the index of
28
refraction and the concentration can be expected (note that Z1
29 p 2 p
this result is, not by accident, similar to the one which is ~vkð~vÞd~v ¼ S ¼ c � S* 2 (7)
30 4 4
obtained from the Lorentz-Lorenz equation for a diluted gas![9]). 0
31
What if the condition c � 1 does not hold? For the index of
32
absorption, it was found that Beer’s law is no longer valid for This integral molar absorbance is equal to the square of the
33
individual spectral points. This finding is closely connected to oscillator strength S2 or the molar oscillator strength S*2 times
34
the properties of the Lorentz-oscillator and the difference the concentration. Equation (7) is one of the so-called Kramers–
35
between it and a Lorentz-profile.[10] The latter is derived from Kronig sum rules. Classically, the oscillator strength was
36
the Lorentz-oscillator by the same assumption that simplifies introduced by dispersion relations, the most prominent of
37
Equation (5) to Equation (6). If Lorentz-profiles really would be which is the classical damped harmonic oscillator model for
38
able to resemble the bands of the absorption index function, which the relative dielectric function is a sum of contributions
39
the bands would be always symmetric around the band from each of the oscillators. It is defined via S2 ¼ q2 N=ðme0 Þ,
40
positions independent of oscillator strength. This would where q is the (partial) charge and μ the reduced mass of the
41
guarantee that absorbance increases linearly with concentra- vibrating atoms. If, for simplicity, we keep assuming only a
42
tion. In contrast, this is not generally the case if Lorentz- single oscillator, the relative dielectric function is given by
43
oscillators are assumed. For higher oscillator strength, the peak [Eq. (8)]:
44
maxima of absorbance bands described by Lorentz-oscillators
45
blueshift relative to the oscillator position (Figure 1, lower S* 2
46 er ð~vÞ ¼ 1 þ c 2 (8)
panel). This blueshift and the corresponding loss of mirror ~v
0 ~v2 i~vy
47
symmetry is due to the fact that the imaginary part of the
48
dielectric function has to be divided by the non-symmetric For wavenumbers clearly below the resonance in the range
49
index of refraction function to obtain the index of absorption where the imaginary part of the relative dielectric function
50
(cf. Figure 1). e00r ð~vÞ � 0, the damping term i~vy is negligible and, since the
51
Still, even in this case, there is a quantity that is preserved wavenumber is lower than the resonance wavenumber ~v0 ,
52
and remains linear to the concentration, which is the integral ~v02 � ~v2 .
53
molar absorbance:[11] Therefore:
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 Communications

1 S* 2
er ð~vÞ � 1 þ c (9)
2 ~v02
3
4
Usually, S* 2 � ~v02 , so that even for large molar concentra-
5
tions, cS* 2 =~v02 �
pffi1ffiffiffiffiin
ffiffiffiffi Equation (9). Accordingly, even for stron-
6
ger oscillators ( cS*2 > 200 cm 1 ),
7
8
S* 2
9 nð~vÞ � 1 þ c (10)
2~v02
10
11
and the increase of the index of refraction due to absorption as
12
implied by Equation (7) can be understood as being conform to
13
the Kramers-Kronig relations (unfortunately a direct prove/
14
derivation seems to be not possible)[11]. In other words, the
15
increase of the index of refraction is proportional to the integral Figure 2. Imaginary part of the relative dielectric function (upper panel), the
16 index of refraction function (center panel, displayed is actually n n1 ) and
absorbance. Nevertheless, the former may be easier to the index of absorption function (lower panel) for different concentrations
17
determine experimentally, at least for strong oscillators, since in between 0.01–0.07 mol L 1 (S*2 = 9801 L/(mol cm2), ~v0 ¼1659 cm 1
18
case of the latter the determination of the baseline may cause γ = 20 cm 1).
19
ambiguities.
20
Note that for real materials we usually have contributions
21
from oscillators at higher wavenumbers (both in the infrared as other hand, the integrated band intensity is, according to
22
well as in the UV-Vis spectral region), the effects of which can Equation (7), a preserved quantity. Even when the integration is
23
be captured by replacing unity in Equation (10) by n1 : carried out only from 1600–1800 cm 1, the increase of the
24
integrated absorbance stays, as already mentioned, to a very
25
S* 2 large degree approximately linear. In practice, however,
26 nð~vÞ ¼ n1 þ c (11)
2~v02 integrating the bands may be problematic, since the result is
27
strongly depending on the choice of the baseline. This is where
28
n1 is the approximately constant index of refraction in the quantitative optical spectroscopy could greatly benefit from
29
transparency region between the infrared and the UV/VIS determining the increase of the index of refraction instead (or
30
spectral region. It is connected to the dielectric background e1 additionally), since this approach is essentially baseline-inde-
31
via e1 ¼ n21 . pendent. Concerning this approach, we compare the increase
32
If we have a mixture or a solution, for which the solvent is of the index of refraction at its maximum for the sample with
33
non-absorbing in the spectral region we are interested in, the the lowest concentration (c = 0.5 mol L 1, 1691 cm 1).
34
solvent might change n1 , the oscillator strength and even the As can be seen in Figure 1, upper panel, this clearly extends
35
oscillator position by chemical interactions, but the linear the range of linearity compared to the case where the
36
change of the index of refraction should be preserved to a pointwise validity of Beer’s law is assumed (orange line in
37
good approximation.[9] Figure 1, upper panel). A further extension of this range is
38
Before we apply the relations on a real system, we first possible, when instead of the maximum of the index of
39
illustrate them employing a model system consisting of one refraction function a preferably strong red-shifted wavenumber
40
Lorentz-oscillator with S*2 = 4900 L / (mol cm2), ~v0 ¼1700 cm 1, is chosen, like, as in Figure 1, upper panel, 1500 cm 1. As
41
γ = 20 cm 1 and c = 50 mol L 1 for the neat substance. For this Equation (10) predicts, the change of the index of refraction at
42
system, for concentrations c < 1 mol L 1 the index of refraction this wavenumber is mostly a linear function of the concen-
43
nð~vÞ stays close to unity (not shown). Therefore, the shape of tration, albeit in a smaller concentration range than the
44
the absorption index band is approximately symmetric like the integrated absorbance. This linear change has been confirmed
45
one for the imaginary part of the relative dielectric function (cf. numerous times experimentally and builds the foundation for
46
lower panel of Figure 2 with upper panel). As soon as the the determination of concentrations by refractive index sensing
47
concentration increases, nð~vÞ begins to significantly differ from and well-established methods like, e. g., sugar concentration
48
unity in the vicinity of the absorption band, as illustrated in determination by refractometer.[12]
49
Figure 1 center panel, while e00r ð~vÞ always stays symmetric For comparably weak oscillators, like most vibrations in
50
(Figure 1, upper panel). organic and biological materials, linearity based on Equation (6)
51
As a consequence, the absorbance band begins to blue-shift could be assumed over the whole concentration range. From
52
with increasing concentration and to show a large tail towards Figure 1, upper panel, we see that linearity holds to about c
53
higher wavenumbers. Accordingly, if we monitor the changes � 8 mol L 1 (comparison of black and pink curve, absorbance
54
of the absorbance at the oscillator position, i. e. at 1700 cm 1, value
p
1
ffiffiffiffiffiffiffiffi at 1700 cm 1). This would be synonymous with
55
we see in Figure 1, upper panel, that for c > 2 mol L 1 the 2
cS* ¼ S ¼ 200 cm , which is approximately the oscillator
56
absorbance increase noticeably deviates from linearity. On the strength of a pure organic or biological material comparable
57

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 Communications

intense C=O vibration, which can be found usually in the range with R2 = 0.99986. Accordingly, not only n n1 , but also its
1
of 1650–1750 cm 1. This limiting value of the oscillator derivative with respect to wavenumber should be linearly
2
strengths also agrees with the limits of applicability of the depending on the concentration. That the slope of Δn around
3
Lorentz-profile, which is based on the same approximation that the inclination/the oscillator position is indeed also a linear
4
leads to Beer’s law, and the need for a Lorentz-oscillator instead function of the concentration is demonstrated in the lower part
5
as shown in Ref. [10]. of Figure 3. This means that at the location of the largest
6
To prove this assumption and to show that Equation (6) is changes, several experimental points can be averaged, which is
7
indeed applicable, we used a quantum cascade laser-based the preferred strategy given that the changes of the index of
8
Mach-Zehnder interferometer which allows to determine refraction are overall quite small. The selection of appropriate
9
absorption and dispersion spectra in the mid-infrared points depends ultimately on spectral resolution and the
10
region.[13,14] With this instrument, we determined the index of damping constant. The latter is to a good approximation the
11
refraction variation of solutions of ɛ-caprolactam in chloroform full width at half height (FWHH). The wavenumbers of the
12
in the region between 1580 and 1730 cm 1, which are due to points at half height are for comparable weak oscillators the
13
the C=O vibration located at 1659 cm 1, in dependence of the positions of the maximum and the minimum of the index of
14
concentration. For the used concentration range (10– refraction. To stay in the linear regime, it is advisable not to
15
70 mmol L 1) comparably small changes of Δn with the wave- take points which are further away than � 1/6 1/8 FWHH from
16
number are expected, and a pointwise linear dependence of Δn the inflection point as a rule of thumb.
17
could be safely assumed. To test this hypothesis, and to The theoretical considerations above are fully consistent
18
illustrate the evaluation of the index of refraction variation with with experimental data. Figure 4 shows the experimentally
19
concentration, we assumed a second model oscillator located at obtained index of refraction functions for solutions of ɛ-
20
1659 cm 1. The further parameters of this oscillator are given by caprolactam in chloroform around the C=O vibration located at
21
S*2 = 9801 L / (mol cm2), γ = 20 cm 1 and c = 9 mol L 1, which is 1659 cm 1 (for experimental details cf. the supporting informa-
22
approximately the concentration of neat ɛ-caprolactam. The tion). Obviously, as already discussed, around the point of
23
corresponding imaginary part of the dielectric function, the
24
index of refraction function and the index of absorption
25
function are displayed in Figure 2.
26
In this concentration range, the oscillator parameters lead
27
to a symmetric shape of the band in the index of absorption
28
function. Correspondingly, the index of refraction function
29
shows a point where n n1 ¼ 0 independent of concentration,
30
which is (almost) located at the oscillator position. At wave-
31
numbers in the vicinity of this point, it seems that the index of
32
refraction changes linearly with wavenumber. This is illustrated
33
in the upper panel of Figure 3. Indeed, linear fits confirm linear
34
variations of Δn with concentration in the range � 2 cm 1
35
around the inflection point ( � 1700 cm 1). Δn, on the other
36
hand, changes linearly with the wavenumber in the same range
37
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54 Figure 3. Upper panel: Δn as a function of concentration for the inflexion Figure 4. Experimentally recorded absorbance (upper panel) and index of
55 point (near zero line) and the points within � 2 cm 1 (one point per 0.5 cm 1, refraction (lower panel) spectra of ɛ-caprolactam at different concentrations.
lines above the zero line refer to lower wavenumbers). Lower panel: Change Inset shows the change of the inclination around the inflexion point as a
56 of the inclination around the inflexion point as a function of concentration. function of concentration.
57

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 Communications

inflection located at the oscillator position the index of Conflict of Interest

1
refraction changes approximately linearly and the inclination
2
changes with the concentration (See inset in Figure 4, lower The authors declare no conflict of interest.
3
panel.) Conform to theory, this change of the inclination with
4
concentration is linear. Keywords: Absorbance · Beer’s law · dispersion · index of
5
In summary, we have extended the theoretical derivation of refraction · vibrational spectroscopy
6
the concentration dependence of the absorbance (“Beer’s law”)
7
to include the concentration dependence of the index of
8
refraction function. For not too strong oscillators in combina-
9
tion with not too high concentrations, the changes of the index
10
of refraction function are linearly related to concentration. This
11 [1] R. Luther, Z. Phys. Chem. 1900, 33U, 245–255.
was demonstrated by experimental results.
12 [2] R. Luther, A. Nikolopulos, Z. Phys. Chem. 1913, 82U, 361–378.
13 [3] H. Kayser, Handbuch der Spektroskopie, Vol. 3, Verlag von S. Hirzel, 1905.
[4] M. Planck, Sitzungsber. K. Preuss. Akad. Wiss. 1903, I, 480–498.
14
Experimental Section [5] T. G. Mayerhöfer, J. Popp, ChemPhysChem 2019, 20, 511–515.
15 [6] T. G. Mayerhöfer, J. Popp, Spectrochim. Acta Part A 2019, 215, 345–347.
16 The experimental section and details about data analysis are [7] T. G. Mayerhöfer, S. Höfer, J. Popp, Phys. Chem. Chem. Phys. 2019, 21,
provided in the Supporting Information. 9793–9801.
17 [8] T. G. Mayerhöfer, J. Popp, J. Opt. A 2006, 8, 657–671.
18 [9] C. C. Wang, J. Y. Tan, L. H. Liu, Appl. Opt. 2017, 56, 7662–7671.
19 [10] T. G. Mayerhöfer, J. Popp, ChemPhysChem 2019, 20, 31–36.
[11] T. G. Mayerhöfer, A. V. Pipa, J. Popp, ChemPhysChem 2019, 20, 2748–
20
Acknowledgements 2753.
21 [12] G. Aronne, P. Malara New Phytol. 2019, 224, 987–993.
22 [13] J. Hayden, S. Hugger, F. Fuchs, B. Lendl Appl. Phys. B. 2018, 124, 29.
Financial support of the EU, the ”Thüringer Ministerium für [14] S. Lindner, J. Hayden, A. Schwaighofer, T. Wolflehner, C. Kristament, M.
23 Gonzales-Cabrera, S. Zlabinger, B. Lendl, Appl. Spectrosc. 2019, 0,
Wirtschaft, Wissenschaft und Digitale Gesellschaft”, the ”Thüringer
24 0003702819892646.
Aufbaubank”, the Federal Ministry of Education and Research,
25
Germany (BMBF), the German Science Foundation, the “Fonds der
26
Chemischen Industrie” and the Carl-Zeiss Foundation is gratefully
27 Manuscript received: January 8, 2020
acknowledged. A.D., A.S., and B.L. gratefully acknowledge funding
28 Revised manuscript received: February 19, 2020
from the European Union’s Horizon 2020 research and innovation Accepted manuscript online: February 19, 2020
29
programme under grant agreement No. 780240. Version of record online: ■■■, ■■■■
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1 COMMUNICATIONS
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Linear or not? Like the absorbance, Dr. T. G. Mayerhöfer*, A. Dabrowska,
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the change of the index of refraction Dr. A. Schwaighofer, Prof. Dr. B. Lendl,
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at a particular spectral point can Prof. Dr. J. Popp
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depend linearly on concentration for
1–6
8 low oscillator strength or concentra-
9 tion. Beyond Beer’s Law: Why the Index
10 of Refraction Depends (Almost)
11 Linearly on Concentration
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