Transcript Hydrophilicity of Pedal Mucus Trails of the Giant African Land Snail Ben Lincoln, Tim Simpson, and Joseph Keddie UniS Department of Physics, University.

Hydrophilicity of Pedal Mucus Trails of the Giant African Land Snail
Ben Lincoln, Tim Simpson, and Joseph Keddie
UniS
Department of Physics, University of Surrey, Guildford, Surrey GU2 7XH, England
Introduction
(a)
Mucus Trails
 Gastropods, such as slugs and snails, secrete a trail of mucus (i.e. a "snail trail") from their pedal
gland while travelling across a surface [1]. The unique mechanical properties of snail pedal mucus
enable the animal's locomotion while also causing the mucus to function as an adhesive to the
substrate [2].
 The mucus trail performs a number of other functions [1], including the provision of mechanisms for
re-tracing a path (i.e. "homing") and for finding a mate of the same species by following a trail [3].
 An understanding of the functionality of trail mucus, including its interactions with water vapour, can
therefore lead to a means of controlling the reproduction of snails and thereby limiting their impact on
the environment, especially vegetable crops [4].
 It has been reported [5] that the presence of snail pedal mucus on a surface influences the
settlement of other adhering marine organisms (such as barnacles) to that surface. The hydrophilicity
or hydrophobicity of a substrate, in particular, has been found to influence the settling of other
organisms [5], which suggests a reason why a mucus trail will have an impact.
(b)
Figure 4. Ellipsometry spectra (y (top) and D
(bottom)) obtained from a mucus film in an
atmosphere with RH = 74.3%. The black line is
the best fit to the data using an EMA model for a
49 nm thick film containing 39 vol.% water.
 The main constituent of gastropod mucus is a complex of proteins and polysaccharides. This
complex is usually classified into the broad categories of mucopolysaccharides and glycoproteins.
 Previous research characterising the "wettability" of mucus has only been semi-quantitative.
Specifically, relative comparisons of hydrophobicity of pedal mucus trails were made via contact-angle
measurements [5].
Infrared spectroscopic ellipsometry (IRSE)
 Infrared spectroscopic ellipsometry (IRSE) is a non-invasive technique that can determine the
structure and optical constants of thin films and bulk materials.
 IRSE has been shown to be a quantitative tool for the measurement of the concentration of
biological molecules in a thin layer [6] and for the determination of the structure of organic films [7].
Objectives
Figure 3.
Simulations to illustrate the
sensitivity of y and D to changes in film
thickness and in water content.
(a)
Simulations of spectra for a film containing 39
vol.% water and having two different
thicknesses: 39 nm (—) and 59 nm (—); (b)
Simulations of spectra for a 49 nm thick film
containing two different water contents: 29
vol.% (—) and 49 vol.% (—).
• To develop IRSE as a technique to detect snail mucus trails in their native state on
a solid substrate.
• To characterise the hydrophilicity of snail pedal mucus by determining the
equilibrium water vapor sorption at constant temperature as a function of the
relative humidity.
Water Content of Film (vol. %)
60
50
40
30
20
10
0
0
20
40
60
80
100
Relative Humidity (%)
Figure 5. Volume fraction of water in the mucus film
as a function of the RH of the atmosphere. The line is
a prediction based on the Flory-Huggins expression
using a best-fit value for the polymer/solvent
interaction parameter of c = 0.54.
Discussion of Results
 There is an extensive literature on the measurement of water vapour sorption in
poly(amino acids), peptides and proteins, as has been reviewed elsewhere [8]. There is a
qualitative similarity between the water vapour sorption of synthetic glassy polymers (such
as poly(vinyl pyrrolidone) and that of biological macromolecules [8]. Accordingly, the
thermodynamic description of solvent sorption that has been developed for synthetic
polymers can, as a first approximation, be applied to natural macromolecules.
 When a polymer is in equilibrium with water in an atmosphere with an activity a, the
volume fraction of water, fw, sorbed in the polymer can be found from
 1
ln a  ln  w  1  1   w   c (1   w ) 2
 N
where c is the Flory-Huggins polymer-solvent interaction parameter and N represents the
number of water molecules that are required to equal the volume of the polymer molecule.
0.04
86
0.035
1000
1500 2000 2500 3000
Wavenumber (cm-1 )
3500
4000
0.025
 A c value of 0.54 0.1 is obtained from the best fit, indicating that the mucus is very
hydrophilic.
0.02
0.015
Conclusions
0.01
0.005
310
0
2000 2200 2400 2600 2800 3000 3200 3400 3600 3800 4000
Wavenumber (cm-1 )
300
290
Δ (Degrees)
280
500
0.03
1000
1500
2000
2500
3000
3500
Amide I
D (degrees)
320
 Water uptake increases strongly when RH increases above 60%. When RH = 84.3%, the
volume fraction of water increases to 53%. This observed trend was reproduced in other
experiments.
CH2, CH3
Ψ
85
Re(D) or Im'(D)
Y (degrees)
87
 Measurements of water sorption in the same mucus film as a function of RH were used to
characterise hydrophilicity, using c as a gauge for comparison to other substances.
Amide N-H
H-bonded OH
IR spectroscopic ellipsometer
Giant African land snail (image
courtesy of Annette Goodman)
84
500
 The parameter c is a gauge of the interaction energy between the polymer and water at a
molecular level. The higher its value, the more hydrophobic is the polymer. A value of c =
0.5 represents a theta-solvent.
0.05
4000
Key Equations for Ellipsometry Analysis:
iD
Ellipticity, r:
r  tanye
where y is related to the change in light’s amplitude
upon reflection, and D is related to the change in
phase.
Optical Density, D:
 ro

D  ln

r
f 

where the “o” superscript refers to the original surface,
and “f” refers to the final (or altered) surface. New
groups on a surface lead to a peak in Re D and a step
with a negative slope in Im D.
C=O
0.025
Amide II
CH2, CH3
0.03
C-O
Figure 1. IR ellipsometry spectra (both y (top) and D
(bottom)) obtained from a bare gold-coated substrate
(
) and from a partially-dried mucus film in the
ambient atmosphere on the same substrate (
).
Re(D) or Im'(D)
0.04
0.035
 The mucus trail remains hydrated under typical ambient conditions. The observed water
sorption is much greater than what has been found in simple proteins (e.g. lysozyme and
bovine serum albumin).
0.02
0.015
0.01
 The hydrophilicity of the mucus might influence the trailing and homing mechanisms of
snails, and it might also provide anti-bacterial protection for the snail.
0.005
0
800
 Characteristic chemical groups for gastropod mucus, including CH2, CH3, CO, C=O, and most importantly - the amide I and II bands, are identified.
 The equilibrium water vapour sorption isotherm of the mucus trail can be described using a
solution model with a Flory-Huggins polymer/solvent interaction parameter of c = 0.54 0.1.
This value is similar to what is obtained for very hydrophilic synthetic polymers, such as
poly(vinyl pyrrolidone).
0.045
Wavenumber (cm-1 )
 Infrared ellipsometry has been used to identify the presence of mucus trails (about 30 nm
thick) deposited by the giant African land snail (Achatina marginata) onto a solid substrate.
1000
1200
1400
1600
1800
2000
Wavenumber (cm-1 )
References
1.
Figure 2. Re D (
) and Im'D (
) spectra for (a) the upper
range of wavenumbers and (b) the lower range of
wavenumbers.
2.
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4.
5.
6.
7.
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