lubricants

Article
Influence of Sample Mixing Techniques on Engine
Oil Contamination Analysis by Infrared Spectroscopy
Torrey Holland 1 , Ali Mazin Abdul-Munaim 2,3 , Dennis G. Watson 2 and
Poopalasingam Sivakumar 1, *
1 Department of Physics, Southern Illinois University Carbondale, 1245 Lincoln Dr. Neckers 483-A,
Carbondale, IL 62901, USA; [email protected]
2 Plant, Soil and Agricultural Systems, Southern Illinois University Carbondale, 1205 Lincoln Dr., Carbondale,
IL 62901, USA; [email protected] (A.M.A.-M.); [email protected] (D.G.W.)
3 Department of Agricultural Machines and Equipment, College of Agricultural Engineering Sciences,
University of Baghdad, Baghdad 10071, Iraq
* Correspondence: [email protected]; Tel.: +1-618-453-5257




Received: 17 November 2018; Accepted: 3 January 2019; Published: 8 January 2019


Abstract: For the most reliable and reproducible results for calibration or general testing purposes
of two immiscible liquids, such as water in engine oil, good emulsification is vital. This study
explores the impact of emulsion quality on the Fourier transform infrared (FT-IR) spectroscopy
calibration standards for measuring water contamination in used or in-service engine oil, in an
attempt to strengthen the specific guidelines of ASTM International standards for sample preparation.
By using different emulsification techniques and readily available laboratory equipment, this work
is an attempt to establish the ideal sample preparation technique for reliability, repeatability, and
reproducibility for FT-IR analysis while still considering the ease and efficiency of the technique.
This study demonstrates that a stable emulsion within a sample, which depends heavily upon the
method, provides a reliably consistent homogenous sample for quantification purposes with FT-IR
analysis. Analysis of variance (ANOVA) modeling and limit of detection calculations demonstrate
the stability of the emulsion. The results reveal that setting a mixing time for a calibration standard
depends on the emulsification process. Inserting a probe directly into a sample (direct sonication)
allows for a rapid, stable emulsion with high reproducibility. Indirect sonication produces relatively
non-miscible liquids of different densities. The pan-shaker produces a reasonably stable emulsion,
but without the long-term stability or quick production time of direct sonication. Reaction time plays
a critical role in the rotary mixing method, which leads to a slow development of emulsification.

Keywords: infrared spectroscopy; mixing methods; emulsion; water contamination; lubricating oils

1. Introduction
To counter the adverse effects of water contamination on the lubricity in an engine, many analytical
methods have been employed to test for contamination to avoid more costly repairs and increased
downtime. When it comes to analyzing water contamination of used or in-service engine oil, FT-IR
analysis stands apart from other commonly employed techniques because it can measure dissolved,
free, and emulsified water, and correlates well with the Karl Fischer method [1–3]. Nuclear magnetic
resonance (NMR) spectroscopy is quite good at detecting all three of these states of water at low
concentrations, but, while it has been employed in research to study the degradation of oil [4] and
even the water content in eggshells [5], it has not yet become a mainstay in commercial labs for
measuring water contamination in engine oil. As NMR devices decrease in cost and size, they may
become more prevalent for this purpose. However, current industry practices show that FT-IR use

Lubricants 2019, 7, 4; doi:10.3390/lubricants7010004 www.mdpi.com/journal/lubricants

Lubricants 2019, 7, 4 2 of 15

is still quite commonplace in this area, and prior research has indicated a need for more specific
guidelines, including within the ASTM International standard of practice for using FT-IR [6], in order
to maximize emulsions in water contaminated oil samples, particularly when preparing samples for
IR calibration [7].
Due to water’s polar nature and the non-polar covalent qualities of oil, they do not readily mix to
any significant degree. Any attraction between water and the hydrocarbon chain of oil molecules is
far less than the affinity between two adjacent water molecules [8]. Additives, however, are mixed
with base oil; engine oil’s additives, in particular the detergents, interact with water and allow
for the formation of relatively stable emulsions. ZDDP (zinc dialkyldithiophosphate), an additive
used for its anti-wear properties, will also increase the oil’s ability to absorb water [9]. Instituting
calibration standards for FT-IR analysis must begin with a thorough emulsification procedure, to
establish an accurate baseline with which to compare an instrument’s capabilities of detecting water
in oil. A thoroughly emulsified sample is inherently more stable, homogeneous, and representative
of the whole, yielding better reproducibility and reliability in results by reducing the variability in
IR measurements that arises due to light scattering in samples with larger water droplet sizes [7,10].
Water contamination in used or in-service oil primarily exists in dissolved or emulsified states due
to agitation, heat, and pressure caused by oil circulation through an engine. Common sampling
procedures also intentionally avoid sampling free water that may exist at the bottom of an engine
oil sump [11]. While the aim of this research primarily focuses on a follow-up to prior research of
calibration samples in FT-IR analysis of engine oil [7], these ideas hold for any sample testing of
immiscible liquids—a representative sample can be better attained through thorough emulsification to
yield homogeneity and reduce variability.
Emulsions have long been studied and employed by the pharmaceutical and food
industries [12,13]. To create a water in oil emulsion without the supplementation of additional
emulsifying agents to the oil matrix, a sufficient amount of energy needs to be supplied to disturb
the oil/water interface such that micro-sized or smaller micelles of water exist within the larger oil
matrix [14]. High-speed stirrers can generate emulsion micelle sizes on the order of a few micrometers,
but obtaining submicron levels in an oil matrix lacking sufficiently high levels of emulsifying agents
requires much higher energy [14,15]. Instability of water in oil emulsions can arise from gravitational
influences on the dispersed phase (water) due to the presence of a density gradient between the
dispersed (water) and continuous phases (oil) [14–16] The process of coalescence leads to sedimentation,
which may be caused by the flocculation of variously sized micelles, or Ostwald ripening, whereby the
larger micelles, being more energetically stable than the smaller ones, scavenge water from smaller
micelles [13–15]. The phase separation caused by the sedimentation process is the driving factor
behind establishing good calibration standards in FT-IR oil analysis, since representative, homogenous
samples are critical in proper enquiries.
In an emulsion, Brownian motion tends to keep the mixture stable if drop size is small enough [15].
A high surface potential can ward off flocculation, while a small droplet size with a negligible
density difference between the dispersed (water) and continuous phases (oil) can prevent creaming or
sedimentation, with creaming having a negative density difference in Stokes’ law:

(ρd − ρc ) gd2
V= (1)
18ηc

where V is the sedimentation or creaming velocity (m/s), ρd is the density of the dispersed phase
(kg/m3 ), ρc is the density of the continuous phase (kg/m3 ), g is the acceleration due to gravity (m/s2 ),
d is the particle diameter (m), and ηc is the dynamic viscosity of the continuous phase (kg/m·s) [16–18].
Equation (1) (Stokes’ law) indicates that micelle diameter plays a major role in the rate of sedimentation
of water in oil, leading to poor calibration standards if mixing does not break up the water droplets
into sufficiently stable sizes.
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Lubricants 2018, 6, x FOR PEER REVIEW 3 of 15

For flocculation
For flocculation to to be
be the
the driving
driving factor
factor in
in emulsion
emulsion instability
instability in aa typical
typical laboratory
laboratory sample,
where the
where the rate
rate of
of flocculation
flocculation isis relative
relative toto Brownian
Brownian motion
motion and
and gravitational
gravitational sedimentation,
sedimentation, an
an
estimate of the ratio of the two types of flocculation rates can be made
estimate of the ratio of the two types of flocculation rates can be made as follows:as follows:
4𝜋(𝜌𝑑 − 𝜌𝑐 )𝑔𝑑𝑖4
𝛤 =4π (ρd − ρc ) gd4i 𝐷𝑖𝑗(1 − 𝐷2𝑖𝑗2) (2)
Γ= 3𝑘𝑇 Dij 1 − Dij (2)
3kT
where 𝛤 is the ratio of sedimentary flocculation to the Brownian flocculation, di is a micelle of one
where Γ is(m),
diameter the kratio of sedimentary
is Boltzmann’s flocculation
constant (m2kg/s2to K),the Brownian
T is flocculation,
the absolute di is in
temperature a micelle
Kelvin,of𝐷𝑖𝑗
one is
diameter (m), k is Boltzmann’s constant 2 2
(m kg/s K), T is the absolute temperature in Kelvin, Dij is the
𝑑𝑖
the ratio of two sizes of micelles ( ), and other parameters are defined as in Equation (1) [17,18]
ratio of two sizes of micelles ddi , 𝑑and 𝑗 other parameters are defined as in Equation (1) [17,18]
Finding the maximum jratio of sedimentary to Brownian flocculation is a matter of
Finding the maximum ratio of sedimentary to Brownian flocculation is a matter of differentiating
differentiating Equation (2) with respect to 𝐷𝑖𝑗 and solving for this size ratio; thus, 𝐷𝑖𝑗√= 𝑑𝑖 ⁄𝑑𝑗 =
Equation (2) with respect to Dij and solving for this size ratio; thus, Dij = di /d j = 1/ 3 ≈ 0.58
1⁄√3 ≈ 0.58
indicates indicates
that the maximum that rate
the of
maximum
sedimentationrate of sedimentation
to Brownian to Brownian
flocculation would flocculation would
occur when micelle
occur when
diameters aremicelle diameters
in a ratio of roughly are3 in
to a5 [17].
ratio Therefore,
of roughlynon-uniformity
3 to 5 [17]. Therefore,
in particlenon-uniformity
size also plays in
a
particle size also plays
factor in phase separations.a factor in phase separations.
Ostwaldripening
Ostwald ripeningcancanalso
alsolead
lead
toto emulsion
emulsion instability,
instability, andand
has has
beenbeen shown
shown to betoreasonably
be reasonably
well
well approximated through the LSW (Lifshitz-Slyozov-Wagner) theory for
approximated through the LSW (Lifshitz-Slyozov-Wagner) theory for emulsions with the equation: emulsions with the
equation:
dr3 3 8DC∞ Vm22γ
ω𝜔== 𝑑𝑟 == 8𝐷𝐶∞ 𝑉𝑚 𝛾 (3)
(3)
dt𝑑𝑡 9RT
9𝑅𝑇
where ω𝜔isisthe
where theOstwald
Ostwaldripening
ripeningraterate(m
(m3 /s),
3/s), rr is
is the
the average
average micelle
micelle radius
radius at at aa given
given time
time (s),
(s), D
D is
is
Einstein’s translational
Einstein’s translational diffusion
diffusion coefficient
coefficientof of the
the dispersed
dispersed phase phase from
from the
the Stokes-Einstein
Stokes-Einstein equation
equation
(m22/s),
(m 𝐶∞
/s), C∞ is
is the
the solubility
solubility of
of the
the dispersed
dispersed phase
phase at
at the
the continuous
continuous phase
phase interface
interface (mol/m
(mol/m 33), 𝑉𝑚
), V is
m is
the molar volume of the dispersed phase micelles (m 33 /mol), 𝛾 is the interfacial
the molar volume of the dispersed phase micelles (m /mol), γ is the interfacial tension between the tension between the
continuousphase
continuous phaseand andthe thedispersed
dispersedphase
phase(J/m(J/m22), andand R R is
is the
the ideal
idealgas
gasconstant
constant(J/mol(J/mol·.K) [13,15,19].
From Equation
From Equation(3) (3)a alinear
linearrelationship
relationship ofof
thethe
cubecube of micelle
of micelle radial
radial sizesize to that
to that of time
of time is implied,
is implied, and
and consequently,
consequently, a stablea stable
emulsionemulsion
shouldshould have amicelle
have a small small sizemicelle
[15].size
Figure[15]. Figure 1 represents
1 represents a graphicala
graphical of
overview overview
emulsion ofinstability,
emulsion instability,
as previously as previously
discussed. discussed.

Figure 1. Routes of instability for emulsified water in oil samples.

Figure 1. Routes of instability for emulsified water in oil samples.

Based on scrutiny of the possible routes of instability, it is implied that a higher rate of energy
Based on scrutiny of the possible routes of instability, it is implied that a higher rate of energy
dispersion throughout the water/oil mixture should induce better emulsification results, generating
dispersion throughout the water/oil mixture should induce better emulsification results, generating
more reliable, readily repeatable results for FT-IR calibration samples with less overall variability.
Lubricants 2019, 7, 4 4 of 15

more reliable, readily repeatable results for FT-IR calibration samples with less overall variability.
This study aims to understand experimentally how the energy dispersion rate throughout the water/oil
mixture affects the emulsion quality, using the different mixing procedures that may be commonly
available with access to modern laboratory equipment to produce better calibration standards for FT-IR
analysis of water in engine oil. The techniques utilized in this study to form emulsions are the rotary
(ROT) method, bath sonication only (BSO), bath sonication followed by a rotary (BSR), pan-shaker
(PSH) method, and direct probe sonication (DPS).

2. Material and Methods

2.1. Sample Preparation

A container (946 mL) of a common diesel engine oil (Shell Rotella T, SAE 15W-40) was purchased
from a domestic retail market in Carbondale, IL, USA. A pipette was used to make different sample
concentrations by volume from fresh oil and non-deionized, distilled water. The treatments included
different water proportions that contaminated the fresh engine oil (0%, 0.1%, 0.2%, 0.5%, 1%, 2%, 5%,
and 10% v/v). A detailed description of the sample preparation used for this study can be found in a
previous publication [7]. In that study, the limit of detection (LOD) was determined, and the aim in
this study was to do a similar determination with similar concentrations. For comparisons of different
mixing methods, the concentrations of 0%, 0.5%, and 1% were used, since concentrations higher than
these tend not to be linear in IR spectroscopy measurements [6].

2.2. Emulsification Procedure

To ensure a sample’s homogeneity for sampling and concentration determination, each sample,
including the control sample, was treated with the same emulsification procedure for each mixing
method. Each sample was stored in a closed vial and left standing upright in a closed box to prevent
photo-degradation. The vial would temporarily be removed for sampling, agitation, FT-IR analysis, or
a photo to track any change in color.
Direct probe sonication of the sample was performed with an ultrasonic processor (Sonic
Vibra-CellTM , model: VCX750, Sonics & Materials Inc., Newtown, CT, USA) with a probe (model:
CV33, Sonics & Materials, Newtown, CT, USA) diameter of 13 mm at a frequency of 20 kHz ± 50 Hz
and power output of 750 W with an amplitude of approximately 76 microns, by inserting the probe
into the vial to within a few millimeters from the bottom. To prevent excessive heating of the sample
during sonication, the vial was placed inside a water bath and the probe was cycled on for 5 s, off
for 30 s. The procedure was repeated for a total of 12 times for a one minute of total sonication time.
The initial and final temperatures of the samples were approximately 22 ◦ C and 40 ◦ C, respectively.
The sonication horn was cleaned with acetone and wiped with extra low-lint Kimtech disposable wipes
after each sample’s agitation. Separate samples that were exposed to direct sonication by this same
apparatus, along with control samples, were sent to a commercial lab for testing of total base number
(TBN) and viscosity, to determine if there was any oxidation of the oil due to incidental heating that
occurred during sonication. No changes were noted. Since each vial sat in a water bath and had to be
opened to allow the insertion of the sonication horn, the opening was shielded from the water bath by
a cellophane tape and paraffin film covering, to prevent possible routes of contamination.
The bath sonicator had a power output of 100 W and a frequency of 42 kHz ± 2.5 kHz. The power
output of the indirect ultrasonic water bath was less than that of the direct ultrasonic probe. Instead
of comparing the power intensity (W/cm2 ) of the two sonicators to estimate the required time for
bath sonication, it was concluded that comparing the sample temperature would be more appropriate.
Ultimately, it was determined that it took 56 min of sonication for the samples in the ultrasonic water
bath to undergo the same temperature change as the direct probe samples had, which was from 22 ◦ C
to 44 ◦ C. The difference was due to the following reasons: first the direct probe method involves direct
contact with the sample and indirectly with the water bath, whereas the bath sonicator is in direct
Lubricants 2019, 7, 4 5 of 15

contact with a larger water bath with greater heat capacitance, and in indirect contact with the sample.
Second, the bath sonicator has a more extensive surface area/air interface with which to dissipate heat
to the atmosphere. The extended sonication time (56 min with an equivalent temperature change) was
chosen to ensure a relatively similar energy absorption by the sample, since bath sonicators can have
cavitational intensities unevenly spread throughout the bath [20]. The power per unit volume was
considerably lower with the bath sonicator, but the hypothesis was that the energy change within the
oil column should have been nearly the same. The vials, sealed with their screw-on caps and paraffin
film, were placed in a beaker of water such that, when submerged, the water level in the beaker rose to
just slightly over the height of the fluid in the vials, yet remained under the bottom of the screw-on
caps. This was to ensure that the entire sample within each vial was exposed to the energy of the
sonicator, but also helped to eliminate another possible path of contamination.
Due to the nature of the bath sonicator driving oscillatory waves within horizontal planes without
vertical mixing or turbulence, as noted by neutrally buoyant particles present in the water bath, it was
hypothesized that an additional mixing method would be required to create a stable, homogeneous
emulsion. A preliminary study was done to establish this necessity. Within this preliminary study only
one sample was used, with two separate replications of each of the four FT-IR measurements, and
these results were compared to the first of the three samples that were ultimately tested using a bath
sonication followed by rotary (BSR) mixing method. Preliminary testing with periodic samplings for
FT-IR analysis indicated that 25 min of additional rotary mixing was sufficient to produce an emulsion,
with an O–H stretching peak that emulated a plateauing of the measurable concentration similar to
that noted in a similar study using a rotary mixer [7]. Therefore, this study includes a bath sonication
method that is followed by 25 min of rotary mixing.
For the rotary mixing method, a continuous rotation was desired to reach full saturation, as noted
by a plateauing of the O–H stretching peak with FT-IR measurements. This was achieved by way of a
Mini Tube Rotator from Fisher Scientific at the maximum setting, 40 rpm. In a preliminary study, it was
established that 60 h was the optimal time to develop full saturation. A similar method was employed
for the pan-shaker method, setting the 15-min mark required to reach full saturation. The pan-shaker
used was a double-arm apparatus (Red Devil Equipment Company, model: Cat 30, Plymouth, MN,
USA) that would accommodate containers with a minimum height of approximately 90 mm. To use
a smaller vial size (27 mm in diameter and 50 mm in height with a total liquid volume of 18 mL),
a wooden spacer with a carved recess specifically made for the vial was used in conjunction with
a metal band that screwed down over the middle of the vial. Equally weighted vials and spacers
were used in each arm. The motion was that of a 30◦ rotation about a central axis at 600 rpm, with a
simultaneous lateral movement from side to side of approximately 5 cm. This double action ensured a
turbulent mixing.

2.3. FT-IR Spectral Analysis

The infrared spectra were recorded with a wavenumber range of 400 to 4000 cm−1 by an
FT-IR spectrometer (Thermo-Nicolet Nexus 670, Thermo Electron Corporation, Madison, WI, USA).
The system was purged with dry air before each measurement (background or sample) to minimize
interference from atmospheric moisture. A background measurement was taken before measuring
each sample. A 3 µL portion of the respective sample was pipetted from the vertical and horizontal
center of the vial onto a KBr window (1 inch diameter), and a similar window was placed atop the
first and rotated 90 degrees to ensure the oil was spread evenly over the windows’ faces. The KBr/oil
sample was placed in a sample tray with a polytetrafluoroethylene (PTFE) ring, sized to ensure the
windows were in the same physical orientation and location in the sample holder each time the
experiment was performed, in order to reduce the possibility of differences in the refraction of light
between each measurement. By gathering the spectra from the same location of the KBr window
during the measurement of the background or sample, spatial dependent spectra were minimized.
After each sample was prepared in a KBr window, four consecutive spectrum measurements were
Lubricants 2019, 7, 4 6 of 15

recorded. The KBr windows were cleaned with methylene chloride after each sample was tested again.
The procedure above was repeated for three different sample preparations of the same concentration.

2.4. Data Preprocessing and Analysis

Before statistical analysis of the mid-peak wavenumber range of the O–H stretching band (from
3150 to 3500 cm−1 ), the baselines of each whole spectrum were adjusted to minimize effects of baseline
shifting between FT-IR measurements. The end of the spectral range [6] (3970 to 3995 cm−1 ), where
each spectrum flattens due to the lack of any detectable signals, was shifted to zero on the absorbance
scale, and the same shift was applied to the entire baseline of each respective spectrum [6].
To determine the concentration of water that could be measured by FT-IR, the peak amplitude
of the entire broad peak water signal around the 3400 cm−1 wavenumbers was used for relative
comparison, using a two-point baseline [21]. The areas of water absorbance were found by two-point
Gaussian curve fitting after eliminating the continuum background, and also by integration from
between the wavenumbers 3150 to 3500 cm−1 . Unless otherwise stated, the integrated values were
used in comparisons.
Water absorbance areas versus water concentration were used to quantify the limit of detection
(LOD) and limit of quantification (LOQ) [22] for water concentrations in oil by FT-IR spectral analysis.
LOD and LOQ were calculated from a linear regression fit, where LOD = 3.3 Sba and LOQ = 10 Sba . Here,
Sa is the standard deviation of the response estimated from y-intercepts of each respective regression
line, and b is the slope.
Significant differences among the data were analyzed using ANOVA (alpha = 0.05) modeling.
When the results were highly significant, p < 0.01 was reported rather than the actual p-value. It was
used to disclose any significant differences among the different mixing methods. This method was
utilized in the areas of the O–H stretch from 3150–3500 cm−1 .

3. Results and Discussion

FT-IR results of bath sonication only (BSO) and bath sonication followed by a rotary (BSR) mixing
were compared with 0%, 0.5%, and 1% water contaminated engine oil, as shown in Figures 2 and 3.
While there were no significant differences (p = 0.18) between BSO and BSR at 0%, the BSR method had
significantly higher values for area in the 3150–3500 cm−1 region than BSO at both 0.5% (p < 0.01) and
1.0% (p < 0.01) rates. At 0.5%, the mean area was 8.1 for BSR, compared to only 2.9 for BSO. At 1.0%,
the mean area was 15.2 for BSR compared to 11 for BSO. When comparing means of contamination
rates for each mixing method, BSO resulted in significant differences (p < 0.01) among the rates but
was only able to distinguish the mean of 1.0% from the lower rates, with no difference between 0%
and 0.5% rates. BSR also resulted in highly significant differences (p < 0.01) among the contamination
rates, with significant differences among each of the rates.
The nature of the wave energy being relatively coplanar with the surface of the water bath of the
bath sonicator did not drive water within the mixing vial into the less dense oil, even though it did
break the water particles apart into particles small enough to allow for emulsification. This is evident
in Figures 2 and 3 of the FT-IR measurements. The horizontal agitation of the mixture when using
a bath sonicator necessitated the use of rotary mixing as well to achieve a full emulsion. Thus, the
BSO method was rejected as a suitable method for generating FT-IR calibration standards of water in
oil emulsions.
An interesting phenomenon was also observed with bath sonication only. After sonication, the
micelles and the small individually dispersed water droplets appeared to maintain stored energy
that could be released with a slight jarring motion by setting down the vials, upon which many of
the micelles would rise geyser-like into the oil column, approaching the upper surface of the oil.
Further mixing was required to disperse the remaining droplets more homogeneously. Figure 4a–d
demonstrates this exciting phenomenon in fluid dynamics.
Lubricants 2018, 6, x FOR PEER REVIEW 7 of 15
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Figure 2. Mean values of the area under the curve (3150–3500 cm− −1) for 0%, 0.5%, and 1%
Figure 2. Mean values of the area under the curve (3150–3500 cm 1 ) for 0%, 0.5%, and 1%
concentrations of water in fresh engine oil for bath sonication only (BSO) and bath sonication plus
Figure 2. Meanof values
concentrations water inoffresh
the engine
area under
oil forthe
bathcurve (3150–3500
sonication cm−1) and
only (BSO) for bath
0%, sonication
0.5%, andplus
1%
rotary mixing (BSR).
concentrations of water in fresh engine oil for bath sonication only (BSO) and bath sonication plus
rotary mixing (BSR).
rotary mixing (BSR).

Figure 3. Bath sonication only of 0%, 0.5%, and 1% water contamination in engine oil, versus bath
Figure 3. Bath sonication only of 0%, 0.5%, and 1% water contamination in engine oil, versus bath
sonication followed by rotary mixing.
sonication followed by rotary mixing.
Figure 3. Bath sonication only of 0%, 0.5%, and 1% water contamination in engine oil, versus bath
sonication followed by rotary mixing.
Lubricants 2019, 7, 4 8 of 15
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a b c d
Figure 4. (a) Water at 1% in fresh engine oil before bath sonication, with two large, distinct water
Figure
droplets4. shown.
(a) WaterForatclarity
1% in vials
fresh depicted
engine oilhere
before
are bath
glass.sonication,
(b) Water with
at 1%two large,engine
in fresh distinct oilwater
after
droplets
bath sonication. A conglomeration of micelles is seen at the bottom of the vial with a small wispbath
shown. For clarity vials depicted here are glass. (b) Water at 1% in fresh engine oil after of a
sonication. A conglomeration
plume (indicated of micelles
by the arrows) is seen
still visible as aatfading
the bottom of thestructure
point-like vial with at a small wisp
the top of amicelle
of the plume
(indicated by theNote
conglomeration. arrows) still visible
the water plume as thata erupted
fading point-like
towards the structure
surface at had the top of the
dissipated micelle
somewhat
conglomeration. Note the
by the time the picture was water
taken.plume that erupted
(c) Water towards
concentration of the
1% surface had
after bath dissipated
sonication somewhat
only, as viewed by
the time down
looking the picture was
into the taken.
vial. From (c)this
Water concentration
perspective, ofappears
the oil 1% afterdarker
bath sonication
in color, andonly,
theasplume
viewed of
looking down into the vial. From this perspective, the oil appears darker in color,
micelles (indicated by the arrows), appearing as the lighter color, can still be seen towards the center and the plume of
micelles (indicated
after they by the
had erupted arrows),
towards theappearing as the
surface after lighter
sitting the color,
vial oncan
thestill be seen
table. towards
(d) Water at 1%theincenter
fresh
after
enginethey
oil had
aftererupted towards
rotary mixing thefollowed
that surface after
bath sitting the vial
sonication. onthe
Note themore
table.homogenous
(d) Water atcolor.
1% in fresh
engine oil after rotary mixing that followed bath sonication. Note the more homogenous color.
In general, the standard deviation appears to be much higher for the BSR method than for the
BSO Inmethod,
general,especially
the standardat the 0.5% concentration.
deviation appears to be Thismuchis,higher
in part, forattributed
the BSR method to the fact
thanthat the
for the
overwhelming
BSO bulk of theatwater
method, especially the 0.5%primarily remained This
concentration. on the is,bottom
in part,ofattributed
the container, andfact
to the wasthat
simply
the
not being sampled
overwhelming bulk with
of theeach
water subsequent
primarily replication
remained on atthe
thebottom
0.5% concentration.
of the container, In addition, only a
and was simply
small
not portion
being sampledof thewith
0.5% or 1.0%
each water concentration
subsequent replication at the erupted
0.5% up into the restInofaddition,
concentration. the oil column;
only a
these portion
small were likely the0.5%
of the smallest
or 1.0%micelles
waterthat were formed
concentration during
erupted uptheintosonication
the rest ofprocedure, and would
the oil column; these
subsequently
were likely the dissipate
smallest into the continuous
micelles that were phase ratherduring
formed homogenously and beprocedure,
the sonication small enough andto would
reduce
the variabilitydissipate
subsequently of the reading
into theduecontinuous
to scatteredphaseinfrared light.homogenously
rather However, when and thisbemass
smallof enough
individual to
water droplets
reduce that remained
the variability of the at the bottom
reading due ofto the vial was
scattered mixed throughout
infrared light. However, the oilwhento getthis
a more
mass truly
of
representative
individual water sample,
dropletsvariably sized micelles
that remained at thewere most
bottom likely
of the vial present,
was mixed and not just the smallest
throughout the oil toones
get
athat hadtruly
more erupted up into thesample,
representative oil. variably sized micelles were most likely present, and not just the
When
smallest ones comparing all mixing
that had erupted up intomethods,
the oil. it was noticed that all are relatively similar to one
another,
When comparing all mixing methods, itdeviations
with much overlap of the standard was noticed ofthat
individual samples and
all are relatively testing
similar procedures.
to one another,
The highest
with variability
much overlap of appeared
the standard in the bath sonication
deviations followed
of individual by rotary
samples and mixing
testing (BSR). This is likely
procedures. The
due to the
highest cavitation
variability being in
appeared non-uniformly spreadfollowed
the bath sonication throughout the water
by rotary mixing bath;
(BSR). a slight
This isvariance
likely duein
location of subsequent samples could lead to poor repeatability in results
to the cavitation being non-uniformly spread throughout the water bath; a slight variance in location [23], which is demonstrated
in Figure
of subsequent5. samples could lead to poor repeatability in results [23], which is demonstrated in
FigureIt 5.
is also worth observing that the highest standard deviations in the O–H stretching band of
FT-IRItresults for individual
is also worth observing samples
that theoccurred in the BSRdeviations
highest standard method, with in thethe standard
O–H stretchingdeviation
band of (STD)
FT-
being
IR 23%for
results of the mean measurement;
individual samples occurred ROT within thethe STD
BSR being 19%
method, withofthe thestandard
mean; PSH with the(STD)
deviation STD
being 23%
18% of the mean; and lastly by DPS,
mean measurement; ROTwithwithSTD beingbeing
the STD 11% of 19% theofmean.
the mean; PSH with the STD
beingThe18%ANOVA results
of the mean; looking
and lastly forby significant
DPS, with STD differences
being 11%betweenof thethe different concentration rates
mean.
according
The ANOVAto mixing method
results are summarized
looking for significantin Table 1. Within
differences each mixing
between method,
the different means of the
concentration 0%,
rates
0.5%, and 1% contamination rates were significantly
according to mixing method are summarized in Table 1. Within different from each other. The rotary mixing
mixing method, means of the (ROT)
method
0%, 0.5%,ofand 1440 1%min, the bath sonication
contamination rates were method of 56 min
significantly followed
different fromby each
rotary mixing
other. Theofrotary
25 min (BSR)
mixing
(ROT) method of 1440 min, the bath sonication method of 56 min followed by rotary mixing of 25
min (BSR) method, the direct probe sonication mixing (DPS) method of 1 min, and the pan shaker
Lubricants 2019, 7, 4 9 of 15

Lubricants 2018, 6, x FOR PEER REVIEW 9 of 15

method, the direct probe sonication mixing (DPS) method of 1 min, and the pan shaker mixing (PSH)
mixing (PSH)
method method
of 15 min of 15 min
all resulted all resulted
in highly in highly
significant significant
differences (p <differences
0.01) among (p the
< 0.01) among the
contamination
contamination
rates, with each rates, with each different
rate significantly rate significantly different
from the others fromrespective
for their the otherstechniques,
for their indicating
respective
techniques, indicating that the mixing times used for each
that the mixing times used for each method were sufficient. method were sufficient.

Figure 5. Areas of the O–H stretching signal (3150–3500 cm−−11 ) for the four different mixing methods
Figure 5. Areas of the O–H stretching signal (3150–3500 cm ) for the four different mixing methods
are seen here with standard deviation error bars.
are seen here with standard deviation error bars.
Table 1. Results for each mixing methods with means of area of 3150–3500 cm−1 wavenumbers of each
Table 1. Results
contamination for each mixing methods with means of area of 3150–3500 cm−1 wavenumbers of each
rate.
contamination rate.
Means of Contamination Rates (a.u.·cm−1 ) *
Mixing Method Means of Contamination Rates (a.u..cm−1)*
Mixing Method 1.0% 0.5% 0%
1.0% 0.5% 0%
Rotary (1440 min) 15.76 8.10 1.33
Rotary (1440 min)
Bath sonication (56 min), then rotary (25 min) 13.78
15.76 8.43
8.10 1.70
1.33
Bath sonication (56 sonication
Direct probe min), then(1rotary
min) (25 min) 16.1313.78 9.10 8.43 1.331.70
DirectPan
probe sonication
shaker (15 min) (1 min) 15.2316.13 10.43 9.10 1.031.33
Pan shaker (15 min) 15.23 10.43 1.03
The four distinct mixing methods were also compared by ANOVA to determine if a significant
The four
difference distinct
existed amongmixing
the methods
methods were also compared
for producing resultsbyatANOVA
the given to concentrations.
determine if a significant
The four
mixing methods were examined at the 0.5% contamination rate, based on the areaThe
difference existed among the methods for producing results at the given concentrations. of four
the
mixing methods
3150–3500 were examined
cm−1 wavenumber at the
range. 0.5%was
There contamination
a significantrate, based on
difference (p <the area
0.05) of thethe
among 3150–3500
mixing
cm−1 wavenumber
methods. The PSH range.
methodTherehad awas a significant
significantly difference
higher absorbance(p < 0.05)
than among
the ROTthe mixingotherwise,
method; methods.
there were no significant differences among the four mixing methods. Based on these results, the were
The PSH method had a significantly higher absorbance than the ROT method; otherwise, there ROT
no significant
method woulddifferences among the Selection
not be recommended. four mixing methods.
among the PSH, Based
DPS,on these
and BSRresults,
methods thewould
ROT method
depend
would
on not be
available recommended.
mixing Selection
time. BSR may amongbased
be rejected the PSH,
on theDPS, and BSRofmethods
requirement would depend
bath sonication and rotaryon
available
mixing mixing time.
equipment, and BSR may be rejected
the extended based on
time required. DPStheisrequirement
preferred forof thebath sonication
shortest mixingand rotary
time, but
mixing equipment, and the extended time required.
if DPS equipment is not available, PSH may be preferred. DPS is preferred for the shortest mixing time,
but ifInDPS equipment
a previous is not
study, available,
each sample PSH may be preferred.
was agitated for about two hours with a rotary device (Mini
In a previous study, each sample was agitated for about two hours with a rotary device (Mini
Tube Rotator, Fisher Scientific, Hampton, NH, USA) and left for about 24 h before performing the
Tube Rotator, Fisher Scientific, Hampton, NH, USA) and left for about 24 h before performing the
FT-IR analysis, to allow any bubbles formed during agitation to disperse [7]. This procedure was
repeated weekly. This study aimed to show the importance of emulsification, and sufficient reaction
Lubricants 2019, 7, 4 10 of 15

FT-IR analysis,
Lubricants 2018, 6, x to
FORallow
PEER any bubbles formed during agitation to disperse [7]. This procedure
REVIEW 10 was
of 15
repeated weekly. This study aimed to show the importance of emulsification, and sufficient reaction
emulsification, in producing
time to form emulsification, producing accurate calibration samples. At week 9 in the previous
study, the
study, the oil
oil appeared
appeared to reach full emulsification, and these results were compared with one minute
direct sonication
of direct sonicationfromfromthis
thisstudy.
study.The
Theresults
resultsofofthe
the0%0%toto10%
10% water
water inin
oiloil concentrations,
concentrations, as as seen
seen in
in Figure
Figure 6, represent
6, represent thethe areas
areas of of
thethe O–H
O–H stretching
stretching from
from 3150–3500cm
3150–3500 − 1
cm after
−1 after one
one minute
minute of direct
probe sonication.
probe sonication. TheseTheseresults
resultsreasonably
reasonablymimic
mimicthe theresults
resultspreviously
previously published
published forfor rotary
rotary mixing
mixing at
at week
week 9 [7].9 [7].

Figure 6. Comparison of FT-IR areas for the O–H stretching (3150–3500 cm−1 ) of water at different
Figure 6. Comparison of FT-IR areas for the O–H stretching (3150–3500 cm−1) of water at different
concentrations after one minute of direct probe sonication.
concentrations after one minute of direct probe sonication.
In order to compare this research with a previous study [7], the peak amplitudes of the first several
In order to
concentrations of compare
water (0%, this research
0.1%, with 1%,
0.2%, 0.5%, a previous
and 2%)study [7], the to
were plotted, peak
use amplitudes of the
linear regression first
fitting
several
to concentrations
establish the LOD and of LOQ
waterby(0%,
the 0.1%, 0.2%,
quickest 0.5%,employed
method 1%, and 2%) were plotted, to use probe
for emulsification—direct linear
regression fitting to establish the LOD and LOQ by the quickest method
sonication. The slope of the first six water concentrations, as mentioned above, were plotted from each employed for
emulsification—direct probe sonication. The slope of the first six water concentrations,
replication, and this was used to calculate the LOD and LOQ for each replication, which were then as mentioned
above, were
averaged. plotted
Figure from the
7 shows eachmean
replication,
values of and thispeak
these was amplitudes
used to calculate
for thethe LOD and
different LOQ for with
replications, each
replication, which were
standard deviation error bars. then averaged. Figure 7 shows the mean values of these peak amplitudes for
the different replications, with standard deviation error bars.
In Figure 8, the results of the LOD and LOQ from a previous study of rotary mixing over
time In
[7]Figure 8, the results
are compared to theofnewtheresults
LOD and LOQ fromprobe
of ultrasonic a previous study
sonication ofof rotary
one mixing
minute totalover time
duration.
[7] are compared to the new results of ultrasonic probe sonication of one minute
The increase in ability to detect lower concentrations of water contamination is evident with direct total duration. The
increase
probe in ability It
sonication. towas
detect lower concentrations
hypothesized of water contamination
that an improvement is evident
in the detection with direct
limit would likely probe
occur
sonication. It was hypothesized that an improvement in the detection limit would
with the use of the ultrasonic probe, as the high-energy sonicator would produce more homogeneity likely occur with by
the use of the
emulsifying theultrasonic
samples to probe, as the
smaller, more high-energy sonicator
uniform micelle would
sizes. More produce
consistent,more homogeneity
smaller, more stable by
emulsifying
micelles the add
should samplesto thetoreproducibility
smaller, more uniform micelle
of tests and sizes.
ensure More
more consistent,
accurate smaller,
sampling when more stable
pipetting.
micelles should add to the reproducibility of tests and ensure more accurate sampling when
pipetting.
Lubricants 2018, 6, x FOR PEER REVIEW 11 of 15
Lubricants 2019, 7, 4 11 of 15
Lubricants 2018, 6, x FOR PEER REVIEW 11 of 15

Figure 7. Peak amplitude of absorbance due to water contamination vs. percent concentration, shown
Figure
with 7. Peak amplitude
standard of absorbance due to water contamination vs. water
percent concentration,inshown
Figure 7. Peak deviation
amplitudeerror bars for 0%,
of absorbance due0.1%, 0.2%,contamination
to water 0.5%, 1%, and vs.
2% percentcontamination fresh
concentration, shown
with standard deviation error bars for 0%, 0.1%, 0.2%, 0.5%, 1%, and 2% water contamination in fresh
engine oil after one minute of sonication from the ultrasonic probe. The dashed line represents
with standard deviation error bars for 0%, 0.1%, 0.2%, 0.5%, 1%, and 2% water contamination in fresh a linear
engine oil after one minute of sonication from the ultrasonic probe. The dashed line represents a
fit.
engine oil after one minute of sonication from the ultrasonic probe. The dashed line represents a linear
linear fit.
fit.

Figure 8. The limit of detection (LOD) and limit of quantification (LOQ) for the percent concentration
Figure 8. in
of water The limit oil
engine of detection
are shown(LOD) andtwo
with for limit of quantification
methods (LOQ) for the percent
of emulsification—rotary mixingconcentration
for two hours
of water
once a in
week engine
for 9 oil are
weeks, shown
and with
one for
minute two
of methods
sonicationof emulsification—rotary
with an ultrasonic probe mixing
[7]. for two hours
Figure 8. The limit of detection (LOD) and limit of quantification (LOQ) for the percent concentration
once a week for 9 weeks, and one minute of sonication with an ultrasonic probe [7].
of water in engine oil are shown with for two methods of emulsification—rotary mixing for two hours
once a week for 9 weeks, and one minute of sonication with an ultrasonic probe [7].
Lubricants 2019,
Lubricants 2018, 7,
6,4x FOR PEER REVIEW 12 of
12 of 15
15

To lend credibility to the hypothesis that using high power output mixing methods to emulsify
To lend
engine oil credibility
and water wouldto thecreate
hypothesis
smallerthat
andusing
morehigh power output
consistently sizedmixing methods
micelles, to emulsify
it is worth noting
engine
that within the four main methods of emulsification, despite having similar FT-IR readings noting
oil and water would create smaller and more consistently sized micelles, it is worth for the
that
samewithin the four main
concentrations, methods
the rotary of emulsification,
mixer (lowest energydespite
outputhaving
per unitsimilar
time)FT-IR
produced readings for the
the highest
same concentrations, the rotary mixer (lowest energy output per unit time) produced
color change (from the golden amber of new, non-contaminated oil to the milkiness seen in the rotary the highest
color
mixedchange
oil and(from thesamples).
water golden amber of new,
Figure 9a–c non-contaminated oil to the
displays this difference in milkiness
color change seenfor in the
therotary
same
mixed oil and water
concentrations samples).
by different Figure
methods. The9a–c displays
micelles this difference
produced after directinprobe
colorsonication
change for thevisible
were same
concentrations by different
but not resolvable with an methods.
optical The micelles produced
microscope with a 100×afterobjective,
direct probeandsonication
using dynamic were visible
light
but not resolvable
scattering (DLS) or with an optical
photon microscope
correlation spectroscopy × objective,
with a 100also and using dynamic
yielded inconclusive resultslight scattering
for likely size
(DLS) or photon correlation spectroscopy also yielded inconclusive results for likely
distribution, likely due to the opacity of the samples tested. They were tested without diluting the size distribution,
likely
samplesdueand
to the opacity of
potentially the samples
altering tested. They
the chemistry, wereproprietary
as the tested without diluting
additives the that
in oil samples and
partially
potentially
dissolve in altering the chemistry,
water could as the proprietary
possibly change additives
their solubility, due toinaoil that partially
change within the dissolve in water
intermolecular
could possibly change their solubility, due to a change within the intermolecular
forces of an altered continuous phase. Conducting a DLS experiment at different concentrations forces of an altered
of a
continuous phase. Conducting a DLS experiment at different concentrations of a nonpolar
nonpolar solvent, and with different solvents, could potentially rule out this effect as negligible or solvent,
and withadifferent
provide solvents,
calibration curve tocould potentially
analyze rule However,
said effect. out this effect as negligible
a more reasonable orapproach
provide awould calibration
be to
curve to analyze said effect. However, a more reasonable approach would be to
use an ultraviolet microscope to more definitively establish the micelle size without disrupting the use an ultraviolet
microscope
chemistry. to more definitively establish the micelle size without disrupting the chemistry.

Figure 9. (a) Fresh engine oil, contaminated with 1% water, before agitation. (b) Fresh engine oil,
Figure 9. (a) Fresh
contaminated engine
with 1% oil,
water, contaminated
after direct probewith 1% water,
sonication. before
(c) Fresh agitation.
engine (b) Fresh engine
oil, contaminated oil,
with 1%
contaminated
water, withmixing.
after rotary 1% water, after direct probe sonication. (c) Fresh engine oil, contaminated with
1% water, after rotary mixing.
As micelles coalesce into larger sizes, the milky color eventually appears in all methods. This milky
appearance showscoalesce
As micelles up in the pan-shaker
into and the
larger sizes, BSRmilky
samples in less
color than anappears
eventually hour, andinin
allthe direct probe
methods. This
samples in severalshows
milky appearance hours,upsuggesting a more stable
in the pan-shaker andsamples
and BSR uniformindistribution
less than anofhour,
nanoemulsion sized
and in the direct
micelles. If nanoemulsions
probe samples are small
in several hours, enough,athey
suggesting morecan be nearly
stable transparent
and uniform [24], but of
distribution thenanoemulsion
driving force
behind emulsion
sized micelles. instability with are
If nanoemulsions suchsmall
nanoemulsions
enough, they is can
Ostwald ripening
be nearly [25,26]. [24],
transparent When butre-sonicated,
the driving
the samples once again lose the stronger milky appearance and return to a
force behind emulsion instability with such nanoemulsions is Ostwald ripening [25,26]. When freshly sonicated color,
re-
which indicates
sonicated, that this once
the samples is notagain
a chemical change
lose the due tomilky
stronger waterappearance
interacting and
with return
oil additives, but a
to a freshly
particle
sonicatedsize change
color, affecting
which the scattering
indicates that this isofnot
light.
a chemical change due to water interacting with oil
additives, but a particle size change affecting the scattering of light.
Lubricants 2019, 7, 4 13 of 15

The ripening of the micelles was timed from the just after sonication to a point in time where they
were resolvable with an optical microscope. With this information, in conjunction with Equation (3),
one can estimate the beginning unresolvable micelle size. Einstein’s translational diffusion coefficient
within Equation (3) can be broken down as follows:

kT
D= (4)
bπr0 ηd

where b is a constant relative to ratios of the diffusing molecules to the continuous phase, r0 is the
initial radius of the diffusing micelles, and ηc is the dynamic viscosity of the continuous phase [27].
Solving numerically for r0 , with C∞ ≈ 24.42 mol/m3 for lubricating oil with additives [28],
Vm ≈ 1.805 × 10−5 m3 /mol, γ ≈ 4.92 × 10−2 J/m2 [29], and ηc ≈ 0.287 kg/m·s [30], the initial micelle
size, ascertained from a directly sonicated sample with 10% water contamination 24 h and 96 h later,
was approximately 140 nm in both instances. The strikingly similar values lend a small bit of credence
to the validity of the equation being able to predict the initial micelle size, and that Ostwald ripening
may be the primary growth factor in this scenario. However, one would expect to be able to resolve
micelles as small as 100–110 nm. These were observable but not resolvable. This discrepancy may
be due to the fact that not all coefficient values for oil were for specifically for 15W-40 Shell Rotella
T. In particular, the solubility of water in the continuous phase C∞ was based on an estimate for
lubricating oil with typical additives. Plus, the solubility estimate is likely low, since the capillary
length of the micelles would be in the order of nanometers [31].
Lastly, it may be of value to illuminate the difference in required mixing time with the rotary
device. A previous study using the same rotary device required approximately 20 total hours of
complete mixing at two-hour intervals over the course of 9 weeks to produce fully emulsified samples
of equal concentrations to those produced under 60 h of constant rotation [7]. This sheds light on the
reaction time it takes for the emulsifiers present within the engine oil to bind with the water to form a
stable emulsion. By drastically increasing the surface area of the water micelles and increasing the
interaction of water binding agents already present in the engine oil through the violent turbulence
caused by direct sonication, this process can be sped up by several orders of magnitude.

4. Conclusions
FT-IR analysis has been employed in the detection of water contamination in engine oil, as it can
detect dissolved water, free water, and water/oil emulsions. A stable emulsion by its very nature
provides a reliably consistent homogenous sample for quantification purposes with FT-IR analysis, yet,
as demonstrated in this study, full emulsification of all free water within a sample depends heavily
upon the device used. As this study suggests, the mixing time used in creating a calibration standard
will vary with the apparatus employed in the emulsification process. The direct application of the
ultrasonic probe proved to be the fastest way to produce a stable emulsion, with less variability
among the samples. Bath sonication has its limitations for the mixing of relatively non-miscible liquids
of different densities, and therefore it is not recommended unless used in conjunction with rotary
mixing to speed up the process if other means are unavailable. The pan-shaker may be an economical
tool to produce emulsions; it may not have the same long-term stability or quick production time
as direct sonication, but can still produce reasonable results. Rotary mixing alone is a rather slow
process for engine oil/water calibration standards, as reaction time plays a critical role. It is also worth
noting that color change alone is not a reliable means of detecting the presence of water in engine oil,
even at relatively high concentration levels, since nanoemulsions can be optically hard to recognize.
In conclusion, the most reliably homogenous calibration samples would be achieved through a direct
probe ultrasonication.

Author Contributions: Conceptualization, T.H., D.G.W. and P.S.; Formal analysis, D.G.W.; Investigation, T.H.,
A.M.A.-M., D.G.W. and P.S.; Methodology, T.H., A.M.A.-M. and P.S.; Resources, P.S.; Software, D.G.W. and P.S.;
Lubricants 2019, 7, 4 14 of 15

Supervision, D.G.W. and P.S.; Validation, A.M.A.-M.; Writing—original draft, T.H., A.M.A.-M., D.G.W. and P.S.;
Writing—review & editing, T.H., D.G.W. and P.S.
Acknowledgments: This research did not receive any specific grant from funding agencies in the public,
commercial, or not-for-profit sectors.
Conflicts of Interest: The authors declare no conflict of interest.

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