Investigation of Paper Chromatography

Flow of Fluid i n Filter Paper

H.4ROLD G . CASSIDY
Sterling C h e m i s t r y L a b o r a t o r y , Y a l e I‘niversity, .Vew Hacen, Conn.

Paper chromatography has become such an important analytical tool that it

seemed desirable to make a fundamental study of the factors that control its
effectiveness. These factors are of three kinds: those which affect flow of liquid,
those which influence RF values, and those which influence definition of zones.
These classes of factors are not mutually exclusive. This paper deals with the
first class of factors, analyzing the factors in terms of bulk properties of paper
and fluid, and of molecular properties of paper and fluid. The major influential
factors of the first kind are porosity of the medium and viscosity of the fluid.
Those of the second kind are more numerous and are examined. The value of
this analysis lies in presenting the active worker with an insight into the com-
plexity of the problem, with a brief survey of what has been done. It serves as a
framework for subsequent reports on the influence of various factors on choice
of solvents for paper chromatography.

S ONE time ago it seemed desirable to begin a fundanieiital

study of the factors which operate in the process of paper
chromatography. The f i s t communication ( 4 0 ) reported the re-
sults of an examination of some 75 filter papers, in which their
suitabilities for the separation of amino acid mixtures were
judged on six counts, using a number of solvent mixtures. It
seemed desirable to report an analysis of the problem, as a frame-
Mork for further reports on experimental findings, and to call at-
tention to a large amount of work which has been done in fields
intimately related to paper chromatography but which, because
it was done with veiy different points of view, is reported in a
variety of jouinals not often available to chromatographers.
In paper chromatography, a developer flows through a paper
strip or sheet, down%-ard(19, 28), upward (35, 47, S Z ) , or hori-
zontally (69). Mediated by the flow of liquid, and oning to the
differential countercurrent nature of the process ( 1 7 ) ,a mixture
of substances may be separated. Paper chromatography re-
sembles chromatography in that the separation de-
pends on a process of distribution between phases which are
brought into differential countercurrent contact. A marked dif-
ference between the two methods is that the former usually re-
lies on capillary forces to set, an “outer” boundary to the moving
phase, while the latter uses a glass, or other, tube as a container.
The developer and mixture may be applied to the paper in one
of several ways. In capillary analysis the strip of paper is dipped
into the mixture, which rises up it, the components being sepa-
rated because some rise less rapidly (are more strongly adsorbed
and retarded) than others. This is the essence of Tiselius’ frontal
analysis (87). The classical capillary analysis method of Schon-
bein ( 7 8 ) and Goppelsroeder ( 9 7 ) is essentially frontal analysis.
In this method there is no development of the chromatogram in
the sense descriged by Tswett (88)and applied by Liesegang ( 4 7 ) ,
and by Consden, Gordon, Martin, and Synge*(19, 28). The
same principle is also utilized when mixtures are analyzed by plac-
ing drops on blotting or filter paper (72). In another method, more
closely related to classical chromatography, the mixture is placed
on the paper as a spot, or streak, and developed chromatographi-
cally with a developer liquid (47). The development may be
carried out by any of the processes known as development analy-
sis (19, 28,47), elution analysis (62, 6 3 ) ,or displacement analysis
(55).
It is an essential part of the method that the fluid developer
move along the paper. Movement of the developer does not
alone guarantee separation, but a sufficient difference in rate of
1415
movement of the components of the mixture must also occur, to-
gether with adequate definition of the zones. There are, there-
fore, a t least three groups of linked factors v-hich operate in the
process of paper chromatography: those which affect the flow of
fluid in the paper, those which affect the Rr values which are ex-
hibited by the components of the mixture, and those which affect
the definition of the zones. This paper deals with the first of
these. A certain arbitrariness is necessary in classifying the vari-
ous factors so as t o make their discussion manageable. TWO sub-
headings have been chosen: the properties of filter paper and the
properties of the fluid. The scheme of discussion is shon-n in
Table I.
BULK-PHASE PROPERTIES OF SUPPORTING MEDlUM
Filter paper is a mat formed by interlaced and to some extent
bonded purified cellulose fibers. The optical microscope shows a
mat structure, and the electron microscope shows (in mechani-
cally disintegrated Whatman KO.0 filter paper) ( 2 ) a brush-heap
structure of what appear to be ribbonlike filaments. Ordinary
filter paper is a simple paper in which the fibers have been matted
to produce a given porosity, and in which the amount of impurity
has been kept under a specified limit. It is this kind of simple
filter paper which is now commonly used in paper chromatog-
raphy, though many other kinds of “papers” of natural and syn-
thetic fibers, glass, and quartz are becoming available.
Two classes of properties of filter paper are of importance t o
this discussion: the properties of the paper as a porous body, and
the properties of the fibers and smaller units, as cellulosic material
(and possibly as porous bodies). These classes need to be distin-
guished because, for example, in speaking of the porosity of filter
paper, as paper, one speaks of a phenomenon very different from
v hat has been termed the “porosity” of cellulose fibers. h dis-
tinction between the roles played by the paper structure and the
fiber structure may not be possible in certain experimental ob-
servations, but this only introduces an added complication which
should be recognized. This distinction appears in Table I as
that betn een bulk properties and niolccular properties.
A convenient way of characterizing the paper as a structure is in
terms of its volumetric composition (1, 49, 60, 62, 7’5): the frac-
tional volumes of solid matter, of air, and of liquid (if any) in a
sheet. The dimensional, total, volume of the paper is determined
by lining up a known number of sheets of the same size and shape
one above the other to form a pad. This is pressed between glass
plates, and the thickness and area of the pad are measured. From
1416 A N A L Y T I C A L CHEMISTRY

Table I. Factors That Operate in Paper Chromatography

I
i I I
Factors that influence Factors that influence Factors that influence
movement of fluid RF value definition of zones

Bulk-phase properties Molecular urooerties

I
Of the paper Of the fluid
1
Of the paper
i

Porosity: Number of cap-
illaries, distribution of
lengths, distribution of
widths, uniformity in prop-
erties
Of the fluid
Type of flow, inertial ef- Chemical nature, presence Chemical atructure as it
fects: in liquid, in displaced of impurities, uniformity influences: density, viscos-
gas of properties ity, surface te sion
I
~

I
~

I
Of the system Of the system
Utilization of driving force Interactions that influence
in overcoming friction, in capillarity, interactions that
changing dimensions, in influence porosity, swelling,
affecting viscosity changes in dimensions,
plugging of capillaries

these measurements and the number of sheets the volume of one
sheet can be calculated.
The fractional volume of solid can be obtained by immersing
the dried evacuated paper in a known volume of a fluid R-hich fills
the pores of the paper but does not swell the fibers, and measur-
ing the increase in volume. The volume of solid fraction may be
difficult to determine because of difficulties also found in deter-
mining the density of the fibers. These have to do with swelling
of the fiber during immersion, complete removal of water, etc.
This problem has been discussed by Hermans and Vermaas (32,
33). The fractional volume of liquid can be obtained approxi-
mately from the loss in weight on drying, and the density of the
volatile material. The liquid fraction which is removed on es-
haustive drying, and which may be largely water, may not actu-
ally represent the fraction that exists as liquid in the paper, but a
portion of it may be a “part” of the fiber in the sense that it is
chemically bound to the fiber ($0). The fractional volume of air
can usually be obtained by difference. Obviously, in making
measurements such as these the size of sample taken must be
large enough that the results no longer depend on the size of the
sample. Only then will one be measuring the properties of the
paper. The attempt to measure these quantities n i t h a sample
consisting of a few fibers would in general yield values not a t all
characteristic of the sheet of paper.
The physical picture one gets a t present of the total filter paper
complex is of a mat of fibers, the pores between which intercon-
nect to form a system of anastomosed capillaries. The fibers
which form the mat themselves contain “pores,” but of submi-
croscopic or even molecular dimensions.
Because of the nature of filter paper it is difficult to give a sharp
picture of the capillary structure: of the number of capillaries and
of their length and width distributions. Instead, the paper is
characterized by various parameters such as “porosity,” “aver-
age,” or “equivalent” pore size, and pore size distribution. These
quantities must not, however, be thought to have necessarily the
same meaning as the same terms used in connection with glass
capillaries, metal tubes and pipes, or even beds of sand.
The entire physical structure of filter paper must be considered
in analyzing the flow of liquids through the paper. Though the
pores in filter paper probably can run in any direction relative to
the surface of the paper, there is none the less not complete ian-
domness of arrangement. (Carman, 16, suggests that in the path
of liquid through granular beds the flow is equivalent to that in a
helically wound capillary, the diameter of the helix being approx-
imately 1.5 times the diameter of the capillary.) The paper
usually differs in structure along the two dimensions of its surface.
These dimensions are defined as the machine direction, parallel to
its forward movement in the paper-making machine, and the
cross direction, a t right angles to the machine direction. The
tIvo are distinguished by tests which rely on: ’
1. Ease of bending. A paper strip cut in the machine direc-
tion tends to bend less easily than one cut in the cross direction.
2. Tendency to curl. If pieces of paper are floated on water,
or, if very absorbent, are exposed to water for a few seconds, the
paper tends to curl. with the axis of curl parallel to the machine
direction (85).
3. Cs illarity. Water usually rises faster up a strip cut in the
machine &-ection. A drop of xater placed on a sheet may spread
to an oval, with the longer avis parallel to the machine direction
(4). (There are some commercial filter papers in which machine
direction cannot be determined by f l o of
~ solvent.)
These properties are consistent uith a preferential lining up of
the fibers in the machine direction as the paper is laid down.
V O L U M E 2 4 , N O . 9, S E P T E M B E R 1 9 5 2
Handmade papers may bho\v increased orientation of the fibers
in one direction. Foote (22) found by application of Lucas’
method (see below) that the calculated pore diameter of several
papers (not filter papers) n-as l a r g c ~in the machine than in the
cross direction.
Manegold ( 5 2 ) reported that a dried filter paper showed a spe-
cific weight (not including the empty space) of 1 3 . The specific
weight’including the empty space was 0.46. After the paper had
&ood 8 days in a desiccator over sulfuric acid its thickness was
1.42 X em., and the yolume of empty space per cubic centi-
meter was 0.71. After 8 days in a water-saturated at.mosphere
the thickness was 1.92 X lo-? em., and the volume of empty
space was 0.79. Thus. swelling increased the thickness and the
fractional volume of empty space in the paper.
Stamm and Millet’t (80)distinguish two kinds of internal sur-
face in filter paper: the eurfaw of the microscopically visible tubu-
lar lumen in natural organized cellulose walls, and of the inter-
fiber spaces; and the surface of “transient capillaries within the
(.ell walls of all types of cellulosic material that exists only in t’he
presence of a swelling agent.” These authors found the area of
the first kind of surface in a filter paper, determined by adsorp-
tion of stearic acid from benzene, to be 2.7 X 103 sq. em. per g r a m
The surface areas obtained in cellulosic systems in which the cell
walls are swollen are of the order of 3 X 106 sy. em. per gram.
BULK PROPERTIES O F THE FLUID
Analyies of the fact,ors influencing rate of flow of fluids (gaws
or liquids) through or in filter paper show, as described in a later
section, that the properties of the fluid which have a major ef-
fect on rate of flow are its viscosity, density, and (Tl-ith liquids)
surface tension. These are niorc conveniently discussed in con-
nection with molecular properties. h further point, of more in-
terest, perhaps, in a discussion of fipI is the observation of Krulla
( $3)that the amount of liquid per unit area of the paper decreases
u p the strip to the advancing front of the liquid.
BULK PROPERTlES OF THE SYSTEM
In measuring the rate of filtration of a liquid through filter
paper ( 7 , 30, Yd, 78, 79) or in general the permeability of a paper
normal to the surface ( 7 5 ) ,a fluid may be forced through the pa-
per under an external pressure, the effect of which is large com-
pared with that of the capillary forces present. Under these
conditions an empirical treatment of the rate of movement of the
fLuid through the paper may be made through the application of
Darcy’s law. This law states that the rate of flow, Q, of a given
liquid through a porous bed is directly proportional to the area of
the bed, A , and the driving pressure, P (the pressure difference
through the porous bed), and inversely proportional t o the thick-
ness of the bed, L. This is stated in the following equation, K
lieing a constant of proportionality:
Q = KAPjL
The equation is valid for laminar (viscous) flow. The constant
Ii is characteristic of the given liquid and porous body. I n the
measurement of rate of filtration, L would be the thickness of t’he
sheet of filter paper, and papers would be compared by using a
given fluid and comparing the values of K so obtained, “without
t~iiquiringinto the size, shape, number, distribution, or orienta-
tion of the actual channels through which the flow takes place”
(36).
\Then Darcy’s law ie applied to the capillary penetration of
liquids into filter paper, it becomes necessary to invoke the theory
of capillarity t.0 define and estimate the driving pressure, P .
One then must inquire into the size, shape, number, and orienta-
tion of the pores in the paper. In applying the theory of capil-
larity, there are also opened up for consideration the molecular
properties oE the fluid as manifested iri surface tension, viscositl-,
and density.
1417
MOLECULAR PROPERTIES OF THE SUPPORTING MEDIUM
An approach similar to that used to the paper as a whole has
been t,aken to the fibers themselves: The fibers also have been
characterized volumetrically as consisting of a certain volume
fraction of cellulose, a fraction of water, and a fraction of empty
space (33). The fibers are very narron- ribbons which seem to be
composed of layers that can be dissected still further ( 6 7 ) .
The intimate structure of the fibers is still under investigation,
but it seems that they show many properties typical of macromo-
lecular systems. Glucose residues are linked together to form
long chains, the cellulose molecules. These are hound together
into larger bodies which eventually constitute the fiber. HOT-
ever, the arrangement appears not to be uniform along the fiber,
but to yield submicroscopic crystalline regions, in which a regular
repeated alignment of the chains is present, alternating with
amorphous regions in n-hich the chains are more or less randomly
kinked and unaligned, the regions shading off the one into thv
other. The empt,y space in t,he fibers may consist of pores of
various sizes, some with dimensions large as compared n-ith the
molecules of common liquids. Into such pores liquids might
penetrate by a process called porous imbibibion.
I n porous imbibition the liquid seems to be taken up by already
existing pores and the porous body does not necessarily change its
dimensions. This is distinguished from swelling, in which p r e
formed pores are not necessarily present, and the process is ac-
companied by dimensional change in the porous body (s8j. 4c-
cording to Hermans ( 3 2 ) , the amorphous parts of the cellulose,
structure are the regions where imbibition leading t o swelling may
occur. The fraction of water which may be present in the fibers
may be made u p of several parts. Some of the moleculeEi nisy be
present in such a form and stoichiometry that they may be con-
sidered as hydrate water molecules. Ot,hers may be more loosely
bound and classified as sorbed miter molecules. Some modern
ideas on this subject have been summarized (76). It is in any case
evident t h a t in filter paper we deal wit’han extremely complicated
system and many variables. It is probable that the volume frac-
tion of water in paper, as measured, is a summation of the water
fraction of the fibers, and of any water which may be held by sorp-
tion between the fibers.
Cellulose. Chemically speaking, t’he solid fraction of filter
paper consists largely of cellulose. This material in filter paper
can usually be separated into three fractions on t’he following
practical basis (45, 84). Alpha-cellulose is insoluble in strong
sodium hydroxide solution. From that part which is soluble,
mineral acid precipitates 6-cellulose. The part which still
remains soluble is ~-cellulose. I t was found that’ a good an-
alytical paper contains at least 96% a-cellulose (86). Reducing
substances are present in the paper, probably largely as part ot
the degraded, soluble fract’ion. These are determined as a copper
number, the number of grams of copper in the cuprous oside
resulting from the reduction of copper sulfate by 100 grams of
paper fibers (85). A good filter paper is found to have a copper
number of 0.4, or less. The acidity of a good paper is found to he
of the order of pH 6 (86).
Cellulose shows a slight solubility in water. Strachan ( 8 1 )
found that a carefully purified cotton cellulose gave up a small
quantity of material, 13 t,o 21 parts per million, even after 50
extractions with water. This material on hydrolysis shoived some
reducing action. Turner (89)found that when a highly purified
cotton fabric was dipped into conductivity water a brown line
formed where the r a t e r evaporated. It could be reformed a t
lower and lower levels on the same piece of fabric with un-
diminished intensity. This indicated some chemical change in
the cellulose lvhich converted it into a water-soluble material.
The brown material was alcohol-soluble and showed reducing
power.
Filter paper cont’ains some “ash,” and many effect’s of its
capillary behavior are laid to the inorganic content (39, 82).
In filter papers which are not *‘filled”with noncellulosic materials,
 1418
the inorganic material may be present as inorganic substances
incorporated in the cellulose by the living plant, as adsorbed salts,
or as cations held by the weakly acid groups present in the paper.
Kullgren (44) reports that in the better grades of filter paper the
ash-forming materials are mostly of the latter type-that is,
cations held through the cation-exchanging ability (largely laid to
carboxylic acid groups) of the cellulose fibers. These cations,
even though present only in traces, may be of considerable im-
portance in chromatography through their catalytic effects
(19).
Some filter papers contain additives introduced for special
purposea. Thus, some papers are treated with melamine or
other substances, to give increased wet strength. Papers are
sometimes filled with filtration aids, such as fuller’s earth or
charcoal. Black papers for use with white precipitates are
made.
Cellulose fibers are swelled by water and many other liquids.
Kress and Bialkowsky ( 4 2 ) ranked liquids in decreasing order
of swelling ability thus: formamide > > water, slightly greater
than ethylene glycol > methyl alcohol > furfuryl alcohol >
ethyl alcohol > furfural, slightly greater than propyl alcohol,
slightly greater than n-butyl alcohol, slightly greater than fuel
oil (a mixture of hydrocarbons slightly heavier than kerosene).
These authors state that liquids with hydroxyl or potential
hydroxyl groups and with high dielectric constants show a high
rate of swelling of cellulose. Liquids with low dielectric constants
did not show much swelling ability. Cellulose is also swollen by
acids and bases within certain concentration ranges. The phe-
nomenon is an exceedingly complicated one, upon which a great
deal of work has been done (29,34,61,71).
FLOW OF FLUID IN FlLTER PAPER
If a strip of filter paper (or any paper which has some porosity)
is dipped into a liquid which wets it, the liquid is observed to
rise up the paper. In the first few seconds after the filter paper
is dipped into the liquid, the liquid seems to enter the paper with
a very turbulent motion (46). The rate of rise, a t first relatively
rapid, slows down gradually as the liquid rises higher in the
paper.
The rise of liquids in filter paper has been studied by a large
number of workers who have been interested in characterizing
the properties of the filter paper, or evaluating the penetrating
power of the liquid, or relating physical t o chemical properties
of the system. Two types of experiments are reported. In one
type of experiment the liquid rises up the paper (or flows down it)
in an enclosed space saturated with the vapors of the liquid.
No evaporation can occur, and the liquid can under these con-
ditions move rather far. The length of ordinary filter paper
sheets limits the observable distances moved, but long rolls
couId be obtained. In packed sand a column of paraffin oil
wm still moving after a year, the height attained being two to
three times that attained in the first 24 hours (31). In the other
type the liquid moves along the paper (blotting or filter paper)
in an open space, so that free evaporation can occur. Under
these circumstances the liquid reaches a certain level a t which
evaporation just balances availability of liquid. The first
type of experiment describes the way in which paper chroma-
tography is usually carried out-that is, in a saturated atmos-
phere.
The earliest studies of the factors which control rate of flow
of liquids in filter or blotting paper were qualitative. The first
quantitative and semitheoretical study seems to have been made
by Cameron and Bell (5,f4)in connection with an investigation of
the movement of soil water. An independent empirical investiga-
tion by Ostwald (60) supported Cameron and Bell. I t remained
for Lucaa (48) in an analysis similar to that of Cameron and Bell,
but more profound, to clarify the relation between height risen,
h, by a liquid in a vertical strip of paper and time of rise, t. He
ANALYTICAL CHEMISTRY
showed that with the aid of the Poiseuille law, made manageable
through suitable assumptions, the relation between h and t
could be derived from theory in the form:
where y is the surface tension of the liquid (assumed t o wet the
paper completely), 7 is the viscosity, and T is a radius of a capillary
which would act in the same way as was observed. On applying
this equation to a large number of data obtained with various
liquids and papers, Lucas found what he felt was good agreement,
considering the simplifying assumptions which had to be made.
For example, the exponents of h fell between 2 and 2.5. The
Lucas relation has been used by paper chemists for estimating the
pore sizes of papers ( 3 6 ) . Lucas realized that his equation im-
plied that the liquid would continue to rise indefinitely. He
showed how this could be corrected for. He also mentioned the
possible effects of sn~elling(see below).
Peek and McLean (64)in connection with work on the im-
pregnation of papers, studied the height of rise of liquids in glass
capillaries and filter paper. Their analysis of the problem was
much like that of Lucas. Recently, very precise measurements
were made by Muller and Clegg (57) on the rise of water, methyl,
ethyl, butyl, and amy1 alcohols, and ethyl alcohol-water mixtures
in three Schleicher and Schull papers Nos. 598-ST and 598-\V
(for spot analysis) and 601. The authors fitted their data with
an equation:
h2 = Dt -b (2)
in which D and b are constants; D is a “diffusion coefficient,”
related to surface tension, viscosity, and density, d of the liquid
by the relation:
D = ay/qd +b (3)
where a and b are constants for a given paper. Here also the
simple equation fits the data over the ordinary range of measure-
ment, so that the equation corrected for maximum height of rise
need not be used.
The expressions which apply Poiseuille’s law are derived from
equations for the f l o ~of liquids through cylindrical capillaries
under known limiting conditions. The exact extension of these
ideas t o filter paper (21, 25, 36, 48, 57, 7 7 ) would require modi-
fications of the theory t o deal with capillaries which branch, and
vary in cross-sectional area and shape. Some such studies have
been made, for example, for capillaries with square, or elliptical,
or in general noncircular cross section (15, 49-51), and for capil-
laries which branch (61-53). Yet even if the mathematics mere
manageable, the extension to filter paper would break down from
a lack of knowledge of the detailed capillary structure of the
paper.
The difficulty was met by Peek and McLean (64) by assuming a
model for the capillary system of the paper. This model “is
one involving a set of capillaries in parallel alternating in series
with larger capillaries, each capillary whether small or large
varying in radius along its length, and therefore similar in its
viscous resistance to a set of shorter capillaries in series.” Each
group of capillaries in series is “replaced by single capillaries
having the same resistance to viscous flow.” The capillaries are
assumed t o vary in size in the model, as in the medium, and,
setting up a distribution function to describe this variation,
Peek and McLean arrive a t an equation which, for practical pur-
poses may be written:
(4)
The derivation is not given because the paper is readily accessi-
ble.
Here “ A and B are dependent only on the distribution of pore
sizes in the medium and are therefore constants for a given
V O L U M E 24, NO. 9, S E P T E M B E R 1 9 5 2
medium.” g is the acceleration due to gravity. The paper strip
is required to have constant width and thickness, in order that
the measurement of the volume of liquid rising per unit time may
reasonably be transformed to a measurement of length, or height,
The plot of the rate of rise dhldt versus l / h should give a straight
line. This was verified a t several temperatures with benzene,
carbon tetrachloride, ethyl alcohol, ethylbenzene, ethylene di-
chloride, and methyl alcohol rising up strips of filter paper in an
atmosphere saturated with the solvent. For a given filter paper
the intercepts of the straight lines obtained with different liquids
should be proportional to d/7, and the values of B (87/dg times
the intercept) should be the same, since they depend only on the
medium and not on the liquid. This also was verified with
various liquids. The slopes of the straight lines are proportional
to ~ / 7 ,and from the slopes relative values of y can be calculated.
A-y is 47 times the slope. Taking a given liquid as having a rela-
tive y of 1, the surface tensions of the other liquids can be cal-
culated. Good agreement was found between calculated and
accepted values of relative surface tension for these liquids.
Peek and McLean showed how a range of pore sizes for the
medium could be estimated, and showed that the estimate for the
filter paper which they used gave a reasonable value.
In all these equations relating h and t, there are tn o classes of
quantities (apart from 9 ) : those which are properties of the
liquid, such as y , d and 7 ; and those which are propeities either
of the capillary, as r is, or of the porous system, as are ,4 and B .
In the latter case, the net effect in fitting the theory derived from
the model to the experimental data is that an adjustable param-
eter is present in the capillary dimensions: the “pore size” or
the “pore size range.”
Quantities r , A, and B . The detailed lengths and cross
sections of the capillaries, or rather the “effective” lengths and
cross sections, since the “length of the capillary” depends upon
the distance from the inlet a t which flow becomes steady, and
the distance from the outlet a t which the flow ceases to be steady
and laminar, are difficult to estimate. However, the front of
rising liquid in a strip of filter paper is usually regular in appear-
ance: the variations iyhich are visible are small, and are averaged
out over-all in the dynamic system. This makes the treatment of
Peek and bicLean applicable, as they point out.
The equation for flow in the capillaries makes no alloxyance for
end effects. I t is k n o m that the shape of the end of a capillary
xi11 affect the rate of flow of a liquid through it. At the entrance
to the capillary there may be a certain distance over r-ihich the
flow of liquid is turbulent. The initial turbulent region is shorter
the smaller the capillary and the less the driving pressure ( 9 I ) ,
so the effect is probably not important with filter paper.
In capillaries of irregular shape the measurement of length
may be open to meniscus errors because the meniscus does not
rise evenly all around its perimeter, particularly if sharp angles, or
cracks, are present (15). This error is of course included in the
general error of not allowing for the mass of liquid above the
bottom of the meniscus in reading h .
Changes in effective radius of the capillaries-that is, in
porosity-might be brought about by the adsorption of colloidal
particles along the ~ - a l l sof the capillaries, or by filtering out of
dispersed material which might block some of the pores, Thus,
it was found (9) that rate of rise of colloid suspensions varied
inversely with the extent of adsorption to the paper of the colloid.
This was influenced by the presence of electrolytes. Entrapment
of air (15,56,18) due to imperfect \vetting (S) may alter porosity.
Bartell and Greager (3)studied the wetting of poxders (which
are porous masses) by liquids and found that when liquids with
large contact angles to the material of the powder were absorbed
(penetrated the powder) air tended to be entrapped. The larger
the contact angle, the more air was entrapped, and the less the
amount of liquid held by the powder. The greatest volume ab-
sorption of liquid was likely to be observed with liquids giving zero
1419
contact angle with the solid, especially if, in addition, the adhesion
tension were low. Liquids with high adhesion tension tended to
draw the particle together, leaving less capillary volume available
for holding liquid. [The adhesion tension A(Ls),was defined
by Freundlich by the relation: A ( L S )= ?(OS) - ~ ( L s ) ,where
) the “surface tension” of the solid, and y ( ~ s
~ ( G s is is)the inter-
facial tension of the liquid-solid interface.] Release of gas by the
liquid ( 7 , 5 6 ) may be another of the causes of change in porosity
when different liquids are compared
The effect of swelling is not sufficiently well understood
(84). It is sometimes assumed ( 2 2 , S O ) that swelling of paper by,
say, water, would tend t o make the capillary spaces smaller.
Holyever, several studies have shown that this is not necessarily
so. Manegold ( 5 2 ) found (the data are given above) that filter
paper after standing in a water-saturated atmosphere was thicker
than the dried paper, and showed about 10% more empty space
volume. In agreement Rith this is the observation of Bogaty
and Carson ( 7 ) ,based on air permeability studies of filter papers,
that the spaces betn-een fibers probably become larger as a result
of swelling in the structure following wetting with water. I t is
not likely that this increased permeability iyould occur only per-
pendicular t o and not parallel to the paper surface. In the swell-
ing process the cellulose fiber gains in thickness, but apparently
not greatly in length.
h sloning of filtration rate is sometimes observed when water
and other fluids are filtered through paper ( 7 , SO, 7 8 ) [or through
glass, porcelain, or quartz filters (78), for that matter]. Pre-
filtering the water usually diminished the slowing ( 7 , 7 8 ) .
The forces due t o change in momentum of the liquid rising in
the capillary are not accounted for in the equation. They
appear t o be very small, especially for capillaries of small radius
(8). Other factors which have been found to play a role in the
flow of fluid through capillaries or porous bodies, are roughness of
the capillary walls ( 5 9 ) ,slippage along the wall ( 4 6 ) , and electro-
osmotic Pack pressure ( 1 1 , I S ) . I t has been suggested ( 7 0 )
that the streaming potential generated in chromatography may
affect the movement of some substances. However, Martin
( 5 4 ) states that this is unlikely, though the zeta potential may
affect the distribution of a charged solute betneen the phases.
These factors may not be very important in influencing the
rise of liquid in filter paper. Many of them must be swallowed
up in the adjustable parameters, A and B , of Equation 4. Some
may operate in opposite ways and so to some extent cancel each
other. It must be evident, however, that filter paper is not a
simple porous body. The walls of the capillaries are flexible,
and capable of s~velling,and the possibility exists that many still
unknown factors are present. If they are, however, it would
seem that their net influence on the flow of the liquid is small.
Quantities y , 7, and d. The liquid would not rise by capil-
larity in the paper if it did not wet the paper to some extent
This, of course, is why there is the y term (replaced by y cos 0
for liquids which make a contact angle e with the paper) in Equa-
tions l, 3, and 4. In the derivation of the equation relating h
and t (Equation 4),it is assumed that the liquid wets the surface
of the capillary with a contact angle of zero. Brown ( I O ) has
pointed out that zero contact angle must not be considered a
special case similar to that represented by a given finite value of
the contact angle (e), but as being characteristic of the class of
circumstances in which when a liquid of surface free energy Y L
wets a solid with the resulting free surface energy Y L ( S ) ,the rela-
tion is:
YL k - YL(S)
What this means is that there is a field of force outside of the
(inextensible) solid surface reaching to the maximum range of
attraction of the solid molecules, and directed inward perpen-
dicularly t o the surface. When a liquid comes into contact with
such a surface, the attraction by the molecules of the solid will
oppose the inward attraction of the molecules in the surface of the
1420
liquid which normally manifests itself in y ~ .The condition for
zero contact angle is that the molecules of the solid attract those
of the liquid to an extent equal to, or greater, than that to which
the liquid molecules attract each other. If the attraction is
greater, then there is a tangential pressure in the surface layer
which tends to cause spreading. If the surface is not inext,ensible,
but, can, for example, be swelled by the liquid, the energy rela-
tions can be changed, but a zero contact angle can still remain
(10).
CONTACT ANGLE. The contact angle of importance in measure-
ments of rate of penetration of liquids is the advancing contact
angle (18); that of a liquid moving on to a fresh surface. (Re-
ceding contact angles, observed on withdrawing liquid from a
wetted surface, appear t o be unstable, and not to have a unique
value.) The presence of pores tends to increase the contact
angle. Thus, surfaces of porous clothing may show an apparent
contact angle of about 150°, when the fibers of which the material
is woven show an angle of about 90" (18). However, where the
contact angle on a material is zero, it is unlikely that mere
macroscopic porosity could increase it to a large value, for a zero
contact angle liquid spreads as a very thin film, probably as a
monolayer, and should easily enter such pores. It seems that
most organic liquids do wet paper in a manner which implies a
zero contact angle (23). Rideal (66) has raised the point that in
measuring the contact angle the alteration of the radius of curva-
ture is observed and measured a t some point distant from the
line of contact with the tube wall, and not at the contact "line"
itself, and he suggests that the angles of wetting (of glass capil-
laries j given in the literature are probably fictitious.
Surface tension may change with change in the velocity of'
flow of the fluid (14). The surface tension of the liquid may
also be altered (usually lowered) on contact with the paper,
either t'hrough the dissolution of surface-active impurit,ies ( 3 7 ,
6 7 ) or through a change in pH. Thus, in contact with solutions
of alkaloidal salts, the filter paper binds acid; this raisp the pH
and alters the degree of hydrolysis of the salt in the direction of
increasing the relative amount of free base. This produces a
fi!ll in surface tension (37); for the free base is more surface
active than the soluble salt. If the liquid wets the paper and
thus penetrates it, the rate will be dependent on viscosity of the
fluid. Thus, if fluids of essentially the same surface tension but
widely different viscosities are allowed to penetrate paper, the
amount of penetration is inversely related to the viscosity. This
was shown by Vincent (go), who used as his penetrants aqueous
polyvinyl acetate solutions.
Viscosity. The presence of the viscosity term in the equations
implies, or is related to, the condition of laminar or viscous
flow in the capillaries of the paper, for if the flow were turbulent
the viscosit,y term \vould disappear. That the flow becomes
laminar after the iiiitial rushing in of liquid is indicated by
analogy to other porouq bodies such as beds of sand (36). The
viscosity of the fluid is Assumed constant, but this assumption
may be invalidated if the liquid shows structural viscosity
change (65). Bulkley (fa)found for some capillaries and fluids
a departure from the Poiseuille law. The viscosity coefficient
may in these cases be a structure-dependent quantity and vary
with rate of flow (51). The assumption may also be invalidated:
if the liquid shows plastic flow, or is thixotropic; if it is a mixture
containing components of different viscosities, one component of
which is adsorbed more strongly than another; if the liquid
contains large molecules which change their average orientation
with change in rate of flow; if the liquid contains charged colloi-
dal particles which are adsorbed along the capillary walls or which
can be discharged under the influence of electromotive forces
generated by the flowing liquid (48);or if the liquid contains
dispersed material (even bacteria, 23) which is filtered out. The
effect of slipping along the walls may be of importance in t.he
contribution to the frictional force due to viscosity ( 4 6 ) .
Density. The density of the liquid enters the equations as
ANALYTICAL CHEMISTRY
part of the term for the force opposing capillary rise: the hydro-
static head of liquid, which is TPLdg. g is a constant, but d
may change if the fluid is a mixture of substances of very different,
densities, one of which is strongly adsorbed. This eventualit,y
is not likely to arise as a serious factor in the application of the
equation.
Cameron and Bell ( 1 4 ) found the same relation between h
and t for blotting paper whether it was in the horizontal or in the
vertical position, which led them to conclude that the capillary
forces were much greater than the gravitational when the dk-
tance risen was small compared to that to which the water might
ultimately rise (penetrate). Berthier (6) ana yzed the forces
operating in the rise of a liquid in a porous body, and the descent
of the liquid. She concluded that only the term for the work
clone against gravity is affected, being positive in the first, and
negative in the second, case. -41~0,if u' is the speed of descent
a t the distance h, and v is the speed of rise at a height h, the func-
+
tion h ( u v') varies linearly with h.
The case where the porous substance dissolves in the liquid
being absorbed was esamined by Russenberger ( 6 8 )using devives
i n which t'he weight of the rising liquid is continually counter-
balanced.
If an esperiment lasts too long when the developer is arid
(or becomes acid for any reason) the behavior of the paper may
be affected. Murray ( 5 8 ) found that adsorption of acid by filter
paper decreases after the paper has been in contact with the arid
for some days. Muller and Clegg ( 6 7 ) found changes in rate of
rise of water in paper on repeated alternating testing followed
by careful drying of the paper.
?VJorsTmE. The effect of moisture has been studied quantita-
tively by Muller and Clegg (57). They found that filter paper
exposed to an atmosphere saturated Tyith water vapor showed an
increased rate of rise with increased time of exposure t,o the
vapors. The maximum rate was found after about 30 minute?'
exposure, though the paper continued to take up water for
several days. This seems to indicate, as a f i s t approximation,
that it is the water of porous imbibition which affects rate of rise
and that any effect as a result of the much slower imbibition
due to s i d l i n g is not marked.
If the paper hangs in a very small quantity of liquid, the lower-
ing in height of the liquid level may affect the rate of rise of the
liquid.
TEMPERATURE. Change in teniperat'uremay affect rate of rise.
Increase in temperature reduces viscosity and surface tension, and
these tend to cancel in their effects on the rate of rise (26).
Lucas calculated the ratio ( y ~ o o / y o o ) / ( q 2 0 0 / ~ ofor
o ) ethyl alcohol
to be 1.4. The ratio should be independent of the porosity of the
paper, and was found for two papers of different porosities to be
1.48 and 1.48 from rate of rise experiments (48). The effect of
temperature has also been investigated by others (57, 6 4 ) with
the conclusion that the temperature coefficient of the rate of rise
is relatively small. The effects calculat,ed when the temperature
coefficient,s of y, q , and d are known, agree with esperiment to
within about 1% in precise work ( 6 7 ) . These conclusions c:~n
be stated only for pure liquids at the present time.
n'hen a volatile liquid is allowed to rise in filter paper in an open
spare so that evaporation may occur freely, the liquid ceases
t'o rise a t that height a t which the amount of liquid rising into
the paper is just balanced by the amount which evaporates.
Garner (25,26)has derived an espression for the effect of evapora-
tion on the rate of climb of liquid under these conditions. An
experimental test indicated that the theoretical equations were
not entirely satisfactory (26). This case is not of importance t o
paper chromatography at present.
Esplicit in the discussion so far has been the requirement' t'h:it
the strips or sheets of paper along which the liquid moves have
constant width and thickness. The shape of a strip will obviously
influence the linear rate of penetration of liquid into it. Thus,
if the point of a triangular strip dips into a liquid which in rising
V O L U M E 2 4 , NO. 9, S E P T E M B E R 1 9 5 2 1421

must spread over a greater and greater area for each unit of
distance risen, even though the available volume may obey the
equations for du/dt (Equation 5), the rate of rise will not obey the
dh/dt equation (Equation 4) derived on the basis that, since
width and thickness remain constant, each increase in volume
will sweep out the same increase in height. Quantitative and
theoretical studies of the effects of shape on rate of rise have been
made by Muller and Clegg ( 5 7 ) . Some effects of shape of the
strip on Rp have been reported ( 4 1 ) .
4CKYOWLEDGMEhT
The author is indebted t o George N. Kowkabany and to
Chester A. Hargreaves, 2nd, for valuable discussions of this
work. He also wishes t o thank the referee for his comments on
commercial filter papers,
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RECEIVED for review December 2, 1951. Accepted June 9, 1952. Commu-
nication S o . I093 from the Sterling Chemistry Laboratory. Report pre-
pared in connection with investigations supported b y a rasearch grant from
the National I n s t i t u t e of Health, Public Health Service