ACTION OF ENZYMES.
“The living organism is enabled by the use of enzymes to bring
about, under ordinary conditions of temperature and moderate
concentrations of acid or alkali, many chemical reactions
which would otherwise require a high temperature or powerful
reagents.”--W. M. BAYLISS.
In a recent work,[94] Dr. Bayliss defines enzymes as the “catalysts
produced by living organisms.” A catalyst is a body which greatly
accelerates the rate of reaction in chemical processes, without
apparently taking part in the process. For instance, peroxide of
hydrogen is decomposed into oxygen and water by mere contact with
finely divided platinum, while the latter remains unaltered in the
process. In this case the platinum black is the catalyst.[95]
[94] “The Nature of Enzyme Action,” by Dr. W. M. Bayliss, FRS.
[95] The important discoveries of MM. Paul Sabatier and Senderens
on the catalytic action of finely divided metals, notably nickel
and copper, have recently been extended to a study of the catalytic
action of various metallic oxides. In the _Comptes rendus_, MM. Paul
Sabatier and A. Mailhe give an account of a new synthetic method,
based on the catalytic effect of titanium oxide, which would appear
to possess many practical applications. They show that if a column
of titanium dioxide is maintained at a temperature of 280°–300° C.,
and a mixture of the vapours of a primary alcohol and a fatty acid
(other than formic acid) is led over it, the corresponding ester
is formed. The same limit is here reached instantaneously as was
found by Berthelot after prolonged contact. An excess of either
constituent favours the limit of combination of the other. Following
this method, the methyl, ethyl, propyl, butyl, isobutyl, and isoamyl
esters of acetic, propionic, butyric, isobutyric, isovaleric and
caproic acids have been prepared. Esters of benzyl alcohol have also
been readily obtained by this method. The inverse action--the direct
hydrolysis of esters by water--is also easily effected, and the use
of titanium dioxide reduces any secondary reactions to a negligible
amount. See Nature, March 9, 1911, p. 54. See also Dr. Sand’s
paper--bibliography.
In natural processes the best known type of an enzyme is diastase
(amylase), the enzyme contained in malt, and which enables the malt
to convert starch into dextrin and sugar (maltose). It is capable of
transforming more than 2000 times its own weight into sugar, which
fact is quite sufficient to show that its action differs from that of
an ordinary chemical reaction. Another enzyme, sucrase, according to
O’Sullivan and Thompson, will hydrolyze 100,000 times its weight of
cane sugar to invert sugar. Rennet will coagulate 250,000 times its
own weight of casein in milk. The list of enzymes grows longer almost
daily, as some new one is separated having a specific action, until
one is almost led to believe that the mechanism of life itself, as
manifested in the cell, is due to enzymes.
It has been found that enzymes act very much in the same way as
inorganic catalysers. As an example, the velocity of the reaction of
invertase (the enzyme of yeast which hydrolyses cane sugar to grape
sugar) has been compared with the same hydrolysis brought about by
heating a solution of cane sugar with a mineral acid. In both cases
the reaction is in accordance with the law of mass action (Guldberg
and Waage) that the amount of sugar transformed will decrease as less
remains to be transformed. In the diagram (Fig. 26) the curve A is for
invertase (Jas. O’Sullivan, “Journ. Inst. of Brewing,” vol. v. p. 168);
curve B is for the hydrolysis by acid (Wilhelmy), from which it will
be seen that the manner in which the hydrolysis proceeds is practically
the same in both cases.[96]
[96] There are apparent exceptions and complications of this law
which we shall not here enter into, except to say that they may be
explained by the fact that the action of some enzymes is reversible
(see p. 141, under lipase.)
[Illustration: Fig. 26.--Curves showing Rate of Hydrolysis.]
In the case of fermentation by the living organism, the fermentation
rises rapidly, and then gradually slows down and comes to an end before
the whole of the fermentable matter is used up; the curve, therefore,
is of a hyperbolic character, C in the diagram, which represents, in a
general way, the fermentation of glucose by B. furfuris. The ordinates
represent the amount of acid produced by the bacteria. The time in this
case would be more nearly represented by hours instead of minutes on
the abscissa.
The mathematical expression for the velocity of the reaction is
_dx_/_dt_ = _k_(_a_ − _x_)
where
_a_ = original concentration of solution
_x_ = the quantity transformed in time t
_k_ = coefficient of velocity of the reaction
By integrating the above equation, it may be shown that
1/_t_ log(_a_/(_a_ − _x_)) = _k_
For the experiment of J. O’Sullivan with invertase, _k_ has a mean
value of 0·0013; for Wilhelmy’s experiment with acid, _k_ has a mean
value of 0·001377.
Enzymes are produced by the living cell, with other secretions, and
as a consequence are found in all plants and animals. Certain organs,
however, produce enzymes in large quantities, or appear to be specially
set apart for their production. In plants, the seeds are the chief
seat of enzyme activity; in animals, certain glands, such as the
salivary glands and the pancreas. The mucous membrane of the stomach
and intestines also secrete enormous quantities.
The production of enzymes by bacteria was observed by Wortmann in
1882. It has been found that the secretion of the enzymes depends
upon the composition of the nutrient medium in which the bacteria are
grown. For instance, Pfeffer found that the secretion of diastase by
Bacillus megatherium depended upon the amount of cane sugar in the
nutrient medium. The cane sugar checked the secretion of the diastase,
and the same effect was observed in the case of the common mould
Penicillium glaucum. In Bacillus mesentericus vulgatus, diastase has
been found to exist side by side with four other enzymes. Passini,[97]
in studying the putrefactive anaerobic bacteria of the normal human
intestines, succeeded in separating from B. putrificus a proteolytic
enzyme filtered free from bacteria, and which caused proteolysis in
media which were too acid to permit of the bacteria growing. The enzyme
easily dissolved, without previous neutralization, the coagulated
casein caused in milk by old coli cultures. Acid gelatin media were
also liquefied by the enzyme.
[97] “Zeit. f. Hygiene,” Bd. xlix. p. 135.
From the evidence we have at present it seems probable that every
variety of enzyme, hydrolysing, oxidizing, ammoniacal, etc., can be
produced by bacteria.
The following useful classification of enzymes is due to Effront.[98]
[98] Les Enzymes, Dr. Jean Effront, Paris, 1899.
----------------------------+-------------------+---------------------
Name of Enzyme |Substance on which | Products of the
| the Enzyme acts | Reaction
----------------------------+-------------------+---------------------
A. HYDROLYZING ENZYMES | |
| |
1. _Fermenting Carbo- | |
hydrates_: | |
Sucrase or invertin |Cane sugar |Invert sugar
Diastase or amylase |Starch and dextrin |Maltose
Maltase or glucase |Dextrin and maltose|Glucose
Lactase |Milk sugar |Glucose and galactose
Trehalase |Trehalose |Glucose
Inulase |Inulin |Levulose
Cytase |Cellulose |Sugars
Pectase |Pectin |Pectates and sugars
Caroubinase |Carobin |Carobinose
| |
2. _Fermenting Glucosides_:| |
Emulsin |Amygdalin and |Glucose, oil of
| other glucosides | bitter almonds,
| | and prussic acid
Myrosin |Myronate of potash |Glucose and allyl
| | isosulphocyanate
Betulase |Gaultherin |Oil of gaultheria,
| | glucose
Rhamnase |Xanthoramin |Rhamnetine,
| | isodulcite
| |
3. _Fermenting Fats_ | |
(Lipolytic): | |
Steapsin |Fats } |Glycerin and
Lipase |Fats } | fatty acids
| |
| |
4. _Fermenting Proteids_: | |
Rennet |Casein |Caseuin
Plasmase |Fibrinogen |Fibrin
Casease |Casein { |
Pepsin |Albuminoids { |Proteoses, peptones
Trypsin} |Ditto { |Proteose, peptones,
Papain } |Ditto { | amides
| |
5. _Fermenting Urea_: | |
Urease |Urea |Ammonium carbonate
| |
B. OXIDIZING FERMENTS | |
Laccase |Uruschic acid, |Oxyuruschic acid,
| tannin, anilin, | various oxidation
| etc. | products
Oxydin |Colouring matters |Ditto
| of cereals |
Malase |Ditto, of fruits |Ditto
Olease |Olive oil |Ditto
Tyrosinase |Tyrosin |Ditto
Oenoxydase |Colouring matter |Ditto
| of wine |
| |
C. FERMENT WHICH SPLITS | |
UP THE MOLECULE | |
Zymase |Various sugars |Alcohol and
| | carbon-dioxide
----------------------------+-------------------+---------------------
A more recent classification based on chemical properties is that of
Kossel and Dakin.[99] They divide ferments into two classes:--
(1) Oxylytic ferments capable of breaking the O-link by which the
radicals are held together in fats and carbohydrates.
(2) Imino-lytic ferments, including the amino-lytic ferments which
act on the amino groups of urea.
Group 2 is sub-divided into--
(_a_) Trypsin and erepsin, which separate the imide NH from the
neighbouring carbonyl CO.
(_b_) Arginase, which separates off urea from arginin.
[99] Zeit. f. Physiol. Chem. xli. f. 153 (1904). See also the
articles by Mr. A. Seymour Jones, B.Sc., writing under the pseudonym
of “Heof Joppa,” in the Leather Trades’ Review, July 19, 1911,
p. 540; and Aug. 16, 1911, p. 625.
Although it is somewhat doubtful whether the enzymes contained in
dog dung are of glandular origin,[100] it is quite certain that other
enzymes are secreted by bacteria developing in the dung while it is
kept prior to being used for puering. These enzymes may be separated
by the following method. About 150 c.c. of puer is well mixed with
an equal quantity of glycerin, and allowed to stand for seven days.
It is then filtered through paper by means of a pump, and yields a
clear filtrate of a deep golden-brown colour; the filtrate is poured
in a thin stream into a tall vessel containing about 1500 c.c. of 98
per cent. alcohol. A copious flocculent precipitate of the albuminous
matter and enzymes is thrown down, the solution is filtered and the
precipitate washed on the filter with absolute alcohol, and then dried
over sulphuric acid in vacuo. The resulting powder is, of course, a
mixture of all the albumins in solution, and probably only a small
portion of it consists of the pure enzymes. We have merely succeeded
by this method in concentrating them. The property of albuminous
bodies in the act of coagulation to carry down soluble matter is well
known, and this also renders the preparation of any pure proteid
extremely difficult. It may be mentioned here that recent evidence
goes to show that enzymes are not of a proteid nature, since, by
repeated purification, the proteid matter may be almost entirely got
rid of, while the activity of the residue containing the enzyme becomes
considerably greater.
[100] See, however, the recent paper by Eberle and Krall, Ueber den
Nachweis des Trypsins im Hundekot, Collegium 1911, p. 201, in which
the authors endeavour to show the presence of unchanged trypsin in
dog dung. Their proof depends upon the action of an antipancreatic
serum on infusions of dog dung based on the work of Achalme, Ann.
de l’Inst. Pasteur, 1901, p. 737. See also the criticism on this
paper by Dr. Otto Röhm and Dr. Max Goldman, Collegium, 1911, p. 265.
Hammarsten (Physiol. Chemistry, 1911, p. 494) states that “among
the secretions which undergo putrefaction in the intestine, the
pancreatic juice, which putrefies most readily, takes place first.”
Krawkov’s method of preparing diastase from saliva (Green, p. 46)
may also be mentioned, as it is of general application. It consists
in salting out the enzymes, by saturating the clear solution (in
this case saliva diluted with an equal volume of water) with neutral
ammonium sulphate. The precipitate which is caused by the saturation is
collected on a filter, and washed for a short time with strong alcohol.
It is then allowed to stand under absolute alcohol for one or two
days, and finally dried at 30° C. On extraction with water it yields
a solution which is strongly diastatic, and which gives no proteid
reactions.
There are several other methods for the preparation and purification of
enzymes, but up to the present it may safely be stated that no one has
succeeded in preparing an enzyme in a state of purity.
In considering the mechanism by which enzymes act, it must be
remembered in the first place that they are colloids, and, as such,
will form absorption compounds with the substrate, or body, upon which
they are acting. It is difficult to understand how an enzyme can
exert an action on the substrate, unless it enters into some kind of
combination with it, although this may be only a temporary one. The
action of some enzymes has been found to be due to extremely small
amounts of certain metals, e.g. in the case of the oxidizing enzyme
laccase, the metal is manganese. In the purest samples of this enzyme
prepared, 0·16 per cent. Mn was found, and it has been supposed by
some observers that the whole of the action of the enzyme may be
attributed to the _physical state_ of the manganese which it contains.
According to this hypothesis,[101] the active part of the enzyme is the
ion Mn. This ion may exist in the solution in two conditions, differing
by the electric charge which they carry. One of them Mn_{++} carries
two positive charges, the other Mn_{+++} carries three. In the first
phase, Mn_{++} is transformed into Mn_{+++} by absorbing the charge of
one ion of hydrogen (H_{+}), and two hydrogen ions thus discharged, in
the _nascent state_, unite with the oxygen dissolved in the liquid to
form water.
[101] Duclaux. See Bibliography.
2Mn_{++} + 2H_{+} + 0 = 2Mn_{+++} + H_{2}O
In the second phase the ion Mn_{+++} with three charges will be
transformed into the ion with two charges, by decomposing a molecule of
water, of which the nascent oxygen will attach itself to the oxidizable
body R yielding the oxide RO.
2Mn_{+++} + H_{2}O + R = 2Mn_{++} + 2H_{+} + RO
and the same cycle of operations will begin again, and continue
indefinitely.
In the above illustration the enzyme action was an oxidizing one.
In the case of puering it is a hydrolytic action, in other words a
molecule of water is added to the skin substance. The fibre or some
portion of it is converted first into proteoses, and finally into
peptones, and simpler bodies. The active metal in these cases appears
to be calcium, but by what mechanism it brings about the hydrolysis is
at present unknown.
Pozerski[102] found that the pancreatic juice, which is secreted
after injections of certain sera (anti-pancreatic action), and as
a consequence has no pancreatic action, contains no calcium, but
pancreatic juice secreted under the influence of pilo-carpin is more
or less rich in calcium, and its proteolytic action increases about
equally with the amount of calcium contained in it. The same probably
holds good for the intestinal juice.
[102] Koch’s Jahresbericht über Gärhungs Organismen, 1908, 635.
Victor Henri has shown that the power of metals in the colloidal state
to bring about these catalytic actions varies with the metal employed,
and is in inverse ratio to the size of the particle. There is a very
interesting and wide field of research open here in order to determine
the conditions under which the various metals act. To this end the
ashes from the purest enzyme preparations might be studied, and methods
devised for producing these metals in the colloidal state, for it seems
evident that it is the _state_ of the body acting which gives it the
properties observed, and not its chemical properties in the usual sense.
The enzymes contained in dog dung which are effective in the puering
process belong to several groups, principally the proteolytic and
lipolytic groups, but indirectly enzymes of the first group (see
p. 132), (fermenting carbohydrates) and of the fifth group (fermenting
urea) also play some part by decomposing various compounds (e.g.
cellulose and urea) contained in the puer.
The action of certain enzymes from the animal body upon skin has
been tried by the author.[103] Those selected were pepsin and
pancreatin,[104] as being most likely to be present in dung. Pepsin
only acts in presence of HCl. Two portions of the same skin were
taken, one of them was treated with a 1 per cent. solution of pepsin,
acidified with 0·2 per cent. of hydrochloric acid; the other in a bate
liquor of dogs’ dung (puer), both at a temperature of 40°C. At the end
of one hour the skin in the pepsin solution was considerably “fallen,”
but that in the manure solution was bated nearly away, i.e. the
greater part of it was dissolved. A 1 per cent. solution of pancreatin
(Mercks) was found to act far more rapidly than pepsin; 1·5 per cent.
of chloroform was added to the solution, to prevent the development of
bacteria. The skin was reduced, but had not the peculiar touch of a
puered skin. As will be shown later, this was found to be due to the
absence of any chemical action upon the lime salts in the skin, and
consequently it felt “limey.” This action took place in the puered
skin, but not in the skin treated with pancreatin alone.
[103] J.S.C.I., vol. xiii., 1894, p. 218.
[104] The word pancreatin is used throughout in the sense of pancreas
extract. As is well known, this contains several enzymes, trypsin,
steapsin, maltase, and also a rennin.
W. J. Salomon (9) has also attributed the activity of the bate to
pepsin and pancreatin, but he does not give any proof of the presence
of these ferments in the bate.
Since it is practically impossible up to the present, to separate these
enzymes from the dung in a state of purity, the method described on
p. 134 was employed.
The enzymes prepared in this way consist of a mixture of all the
enzymes present in the dung, the amount obtained from 1000 grm. of
dung being about 4 grm. The product had a slight diastatic action upon
starch; 0·5 grm. in 100 c.c. of water at 35° C. was found to have a
very considerable reducing action upon skin, and when combined with the
amine compounds prepared from the dung, the action was more powerful,
and more rapid than with puer. Limed skin was puered in thirty minutes
in this solution to a perfect condition, in the absence of bacteria,
and with no evil smelling compounds. The reaction of the solution at
the beginning of the experiment was faintly alkaline; at the end of the
experiment it was considerably alkaline.
This experiment proves that the action of the dung is a complex one,
due to the combined action of enzymes and chemical compounds upon the
skin. These compounds, which are principally amines, and salts of
amino-acids, probably assist the enzymes, and at the same time act
upon the lime remaining in the skin from the previous liming process.
Whether a skin, which has never been submitted to the liming operation,
could be bated by enzymes alone, without the addition of amines, has
not, so far as I know, been tried, but it is highly probable that this
would be the case.
In order to compare the action of the enzymes prepared from dung with
that of the enzymes produced by bacteria, a mixed culture of bacteria
from puer in dextrose gelatin, after seven days’ growth, was taken.
200 c.c. of this was mixed with 200 c.c. of dilute alcohol (65 per
cent.), and well shaken: gelatin and albuminoid bodies are by this
means precipitated. The liquid was filtered and poured into eight times
its volume of 98 per cent. alcohol. The precipitate which came down
was washed with absolute alcohol, and dried in vacuo. The enzymes thus
obtained were re-dissolved in water, and the former experiment with
the skin repeated with this solution, with the addition of the amines.
The skin was brought down in exactly the same way as before, showing
conclusively that it is the enzymes produced by bacteria, acting in
conjunction with the amines, which bate the skin. It would seem that
the action of the enzymes is aided by the presence of amine compounds,
in addition to the chemical action which these latter have upon the
skin. The action is interdependent, i.e. bacterial action alone is
insufficient, and chemical action alone is insufficient, the true
bating action being a combination of the two.
There seems little doubt that it is the enzymes which dissolve the skin
substance, or rather certain parts of the intercellular substance of
the fibres, and the compound of this substance with lime. The action is
a digestive one, and may be compared, as we have shown, to that of the
digestive ferment of the pancreas.
This fact has been made use of in the artificial bate “Oropon” (see
Chapter VII.), in which extract of pancreas is combined with ammonium
chloride, and some inert material.
The effect of puering on the fatty matter in the skin is well known.
The fat and grease are partially emulsified, and set free, so that
they can be removed by scudding. This action is a most important one,
and one in which artificial bates have hitherto been wanting. The
emulsification of the fats is brought about by means of an enzyme
either identical with, or closely resembling, lipase.[105] This enzyme
is found in the pancreatic juice, and in the seeds of many plants. It
brings about the emulsification of the fat by saponifying a portion
of it, i.e. the fat is split up into glycerin[106] and a fatty acid,
according to the equation--
C_{3}H_{5}(C_{18}H_{35}O_{2})_{3} + 3H_{2}O
Stearin.
= C_{3}H_{5}(OH)_{3} + 3(C_{18}H_{35}OHO)
Glycerin. Stearic Acid.
[105] Also known as steapsin, or pyolin (Allen, p. 357, vol. iv.,
Comm. Org. Analysis).
[106] In the digestion of fat by dogs the glycerol produced is all
absorbed before the ileum is reached, so that none exists in the
excrement. (Levites, Chem. Soc. Abst. 1907, vol. iv. p. 891.)
Lipase was one of the first enzymes in which the reversibility of
the reaction was shown, i.e. it is not only capable of hydrolysing a
fat, but also of causing the formation of one by the combination of
the fatty acid and glycerin.[107] This explains why the reaction of
such an enzyme is never complete. An equilibrium is reached just as
with ordinary reversible chemical reactions like the precipitation of
magnesium hydroxide by ammonia.
[107] Another instance of the reversibility of enzyme action is the
secretion of a peptolytic enzyme, by B. pyocyaneus; i.e. it has the
power of synthesizing proteins, as well as of decomposing them. (Zak.
Chem. Soc. Abst. 1907, p. 996.)
Lecithin, and possibly other fatty compounds, are known[108] to be
important auxiliaries in the ferment-like actions produced by toxins;
cholesterol has a similar effect, and as this body is a constituent
of dung, it may play some part in puering. Here, again, is a further
problem awaiting investigation.
[108] Chem. Soc. Annual Reports IV., 1907, p. 252.
Loevenhart has shown that the bile salts, sodium cholate, and sodium
glycocholate, greatly increase the activity of lipase, and Magnus found
that synthetic bile salts have the same effect; such bodies are known
as co-enzymes. I have shown that bile itself is not favourable to the
bating action, but the bile salts, by stimulating the action of the fat
splitting enzymes, are probably essential to the full action of the
dung bate.
Another enzyme which may be of importance in puering is erepsin, the
enzyme of the intestinal juice, which is responsible for the completion
of the digestive process. The pancreatic enzymes act upon the peptones
produced by the pepsin of the stomach, splitting them up into simpler
compounds, while the erepsin acts further upon these products. It
dissociates albumoses and peptones into amino acids, taking as it
were the last traces of nutriment from the food passing through the
intestine. It acts best in alkaline solutions.
This ferment is very widely distributed in the animal kingdom, and
occurs in other organs and tissues besides the intestines. The quantity
of erepsin in the fresh fæces must be considerable, since a dog
secretes from 400 to 500 c.c. of intestinal juice per day. It remains
to be shown whether this retains its properties after excretion, and
for how long.
A most important point in connexion with the activity of enzymes is
the reaction of the medium, i.e. its acidity or alkalinity, or, more
strictly speaking, its hydrogen ion concentration. A very slight
increase or decrease of the acidity or alkalinity of the liquor will
diminish the rate of action of the enzyme by a large amount, and in
some cases cause the action to cease. In all cases enzymes have an
optimum acidity or alkalinity; in other words, for every enzyme there
is a particular hydrogen ion concentration at which its activity is at
a maximum. The work of Soerensen, to which I have already referred,
gives a very complete account of this aspect of enzyme action and also
of the methods he employed to investigate it. His work should certainly
be carefully studied by anyone wishing to take up this part of the
subject.
The same remarks as to optimum conditions apply to temperature,
although the effect of this is better known. Most tryptic enzymes act
best at the body temperature, viz. 98° to 100° F. (37° to 40° C.), and
hence puering should be conducted at this temperature. In the case of
the hen or pigeon-dung bate the enzymes have not yet been studied, so
far as I am aware, so that it is not possible to give an account of
them. At the same time it seems highly probable that, as these bates
are employed at comparatively low temperatures, the enzyme action is
kept back, and therefore the chief action would be a chemical one.
Enzymes are retarded in their action in the first place by their own
products, in a very similar manner to bacteria, in the second place
the retarding or inhibiting action is brought about by the so-called
_anti-bodies_. Of these the longest known and best studied are those
which are produced by toxins, and which neutralize the action of the
toxins upon the animal organism (anti-toxins).[109] Normal serum also
inhibits the action of trypsin and many other enzymes. Another very
important group of anti-bodies are the _precipitins_. If the serum
of an animal be injected repeatedly into another animal of different
species, a precipitin appears in the serum of the animal treated, which
causes a precipitate when added to the serum of the first animal. The
special importance of this fact is, that it can be utilized as a method
of distinguishing between human blood and that of animals, which is
often of importance in medical juris-prudence.[110]
[109] Hammarsten, p. 70.
[110] Gulland, Ency. Britt. iv., p. 83.
This fact has been made use of to distinguish dog dung from other
matters, with which it has been adulterated.[111] A perfectly clear
extract of dog dung filtered free from bacteria was injected into a
rabbit. The serum obtained from the rabbit was found to contain a
precipitin, and on addition of the serum to the dog dung extract a
precipitate was produced. When the serum was added to the extract from
the dung of another animal no precipitate was formed. When added to the
extract of the adulterated dung a very much smaller precipitate was
produced than with the extract from dog dung alone.
[111] Appelius, Technische Briefe, No. 23, April 1909.
The coli bacteria in the puer also produce an
anti-body--_agglutinin_.[112] If a culture of B. coli be examined under
the microscope, the bacteria are seen moving here and there in the
liquid, evenly distributed. On the addition of a trace of the serum
of an animal which has been previously injected with coli bacteria
the bacteria on the slide cease their movements and collect together
in clumps. They are said to be agglutinated. This property is used to
diagnose bacteria in suspected cases of cholera and some other diseases.
[112] Harden, see Bibliography.
The wonderful discoveries that have been made in this direction
constitute one of the most marvellous chapters in the history of
science. Serum diagnosis and serum therapeutics are now firmly
established as invaluable aids to the physician in his fight against
disease and death.
The extent of our present knowledge of the action of enzymes in puering
may be summed up as follows. Active enzymes are produced by bacteria
growing in the infusion of dung, in addition to digestive enzymes
which may be originally present in the dung; the bacterial enzymes
are produced more rapidly in a dilute infusion, as employed in the
puer wheel, than in the dung itself. The enzymes are of various kinds,
proteolytic, peptolytic, lipolytic, etc., but the proteolytic and
lipolytic are the most important. These have a solvent action on the
fibres of the skin, but little or no action on the hyaline layer, at
the concentration usually found in the puer liquor. The fatty matters
and soaps in the skin are acted upon by the lipolytic enzymes, and the
fats to some extent emulsified, so that they may be easily removed from
the skin by scudding or pressing.
It must be clearly understood that enzyme action alone is not
sufficient, as has been previously explained, but that the dung
enzymes, acting in conjunction with the chemical compounds present,
produce the specific puering effect.
There is still much work to be done before the action of the enzymes in
dung is fully understood, but from the above short account it will be
seen that the part they play in the bate is of great importance.