NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH CATTLE
229
be gauged for the present purpose by comparing the amounts ingested with the
amounts excreted in the urine.
Four sets of determinations were made. In the first three tests the sulphate in-
gested was simply the amount present in the straw. In the fourth test additional
sulphate was fed. The results summarised in Table III show that invariably a loss
of ingested sulphate takes place.
TABLE III.
Sulphate balance experiments with the ragi straw ration.
Expt.
1
2
3
4
Na 2 SO 4
Supplement
0
0
0
6
Sulphate in grms. sulphur per day
G
H
G H
G
H
G
H
Intake
. 5.894
. 5.920
. 5.555
. 6.144
. 4.909
. 5.504
. 5.808
. 6.404
Outgo
4.531
4.599
4.366
4.646
3.911
4.184
4.576
5.040
Loss
1.163
1.320
1.189
1.498
0.998
1.324
1.232
1.364
(c) Total sulphur balance.
Two series of tests were carried out. The results are shown in Table IV.
The figures agree generally in showing that the intake and outgo are fairly well
accounted for. No stress is laid on the final balance. It should be remarked,
however, that the results are corroborated in a qualitative sense by the fact that
the favourable balances of the first test coincide with a period during which the
animals were eating well, while the negative balances were obtained at a time when
the animals were somewhat off their food.
230 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, III
TABLE IV.
Sulphur balance experiments with ragi straw.
(Sulphate and organic sulphur as grms. S per day).
—
Intake—
Total Sulphur
.
.
.
.
.
Sulphate
Intake—
Organic Sulphur
Outgo—
Total Sulphur
.
.
.
.
.
Sulphate
Outgo—
Organic Sulphur
Increase of Organic Sulphur . . .
Decrease of Inorganic Sulphate . . .
Sulphur Balance
.
.
.
.
.
First test
9.858
5.952
3.906
9.258
4.361
4.897
0.991
1.591
+0.600
10.138
6.109
4.029
9.662
4.650
5.012
0.983
1.459
+ 0.476
Second test
7.642
4.903
2.739
7.968
3.911
4.057
1.318
0.992
—0.326
8.321
5.361
2.960
8.524
4.180
4.344
1.384
1.181
—0.203
It is clear that loss of sulphate occurs. It is equally clear that this loss of
sulphate is not accompanied by a parallel loss of sulphur. The data are an indica-
tion of what is taking place. More accurate figures, obtained under exceptionally
favourable conditions, are given in a later table.
EXPERIMENTS WITH MATURE HAY.
(a) Excretion of volatile phenols and sulphates.
Simultaneous determinations of volatile phenols and sulphates were made
firstly on the basal ration and secondly on the ration supplemented with sodium
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH CATTLE
231
sulphate. Further determinations of sulphates only were then made after incre-
ment of the sulphate supplement. The results are shown in the accompanying
Table.
TABLE V.
Urinary excretion of sulphates and phenols with the hay ration.
(Sulphate and phenol expressed as grms. S per day).
Test
1
2
3
4
6
Grms.
Na 2 SO 4
Supplement
0
6
6
9
15
Total SO 4
G 1.546
H 1.889
G 2.788
H 2.750
G 2.516
H 2.752
G 3.062
H 3.081
G 4.275
H 4.446
Free SO 4
0
0
0.422
0.512
0.155
0.497
0.789
0.889
2.016
2.026
Ethereal SO 4
1.546
1.889
2.367
2.237
2.361
2.256
2.273
2.192
2.260
2.421
Volatile
Phenol
1.380
1.700
1.609
1.542
..
..
..
..
..
..
Other
ethereal
SO 4
..
..
0.758
0.695
..
..
..
..
..
..
In the first test the sulphate and phenol excretion of H is somewhat above the
average for this animal. 1.5 to 1.6 is a fair daily figure for both the animals.
The sulphate figures in the first test require attention. It will be noticed that
this urine did not contain any free sulphate, the entire amount was present in the
ethereal form. In this case, where there is obvious shortage of sulphur, the ethereal
sulphates corresponds closely to the amount of volatile phenol (p. cresol). In the
second test, the points to note are, firstly, the presence of a small amount of free
sulphate, secondly, a very considerable increase of ethereal sulphate, which exceeds
the volatile phenol determined simultaneously.
In this urine, therefore, there are bodies present, other than p. cresol. which
conjugate preferentially with sulphate ; but their avidity for this conjugation is not
sufficiently intense to enable them to procure sulphate when there is a shortage of
sulphur. The data emphasise the avidity of p. cresol which apparently conjugated
completely with sulphate when there was a decided deficiency of sulphur.
Tests 3, 4 and 5 show that when excess of sulphate is present, the ethereal
sulphates attain a maximum constant which is not affected by greatly increasing
the excess of sulphate. The ethereal sulphate is evidently a measure of the total
phenolic bodies present in the urine. The volatile phenol being known, the
232 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, III
difference represents other phenolic substances, which remain remarkably constant
in amount, being equivalent to .7 grm. S.
The hay urine differs in this respect, in a marked manner from ragi straw urine,
in which the phenols were shown above to consist almost entirely of p. cresol.
The characteristics of the hay urine may by summarised thus :—
1. A deficiency of sulphur is indicated by low values for total sulphate and
complete absence of free sulphate.
2. The ethereal sulphate is equivalent to the volatile phenol which seems
to indicate a very strong preference of p. cresol for the sulphuric
conjugation.
3. Appreciable quantities of other phenolic bodies are present. These also
have a preference for the sulphuric acid conjugation, which, however,
cannot take place unless an excess of sulphate is provided.
(b) Sulphate balance experiments.
Four sets of figures for the sulphate balance were procured, the first test being
made on the hay ration without sulphate supplement, the succeeding tests with
increasing quantities of Na 2 SO 4 as supplement. The results obtained are shown in
Table VI in which it is seen that loss of sulphate invariably takes place with this
ration.
TABLE VI.
Sulphate balance experiments with mature hay.
Experiment
No.
1
2
3
4
Grms. Na 2 SO 4
Supplement
0
6
9
15
Sulphate as grms. S per day
Intake
G
H
G
H
G
H
G
H
.
.
.
.
.
.
.
.
2.211
2.289
3.481
3.628
4.315
4.523
5.715
5.875
Outgo
1.708
1.728
2.517
2.727
3.126
3.178
4.362
1.368
Loss
0.503
0.561
0.964
0.901
1.189
1.345
1.353
1.507
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH CATTLE
233
It should be pointed out that the sulphate in the faeces has been neglected in
calculating these balances. Corrected values, however, are shown in a subsequent
statement.
(c) Total sulphur balances.
In order to obtain a striking contrast, two tests, one without a sulphate sup-
plement, and one with the maximum sulphate dosage, were selected for this work.
Further, to attain the greatest possible analytical accuracy, the corresponding
samples from the two tests were analysed simultaneously in every case—for
example, the two samples of hay were oxidised side by side and the entire process
carried through simultaneously with the two samples which were to be contrasted.
The sulphur balance has been calculated with the aid of these figures, the
digestion data and the corresponding urine analyses. The results obtained are
shown in the following table :—
TABLE VII.
Sulphur balance experiments with the hay ration.
(All forms of sulphur expressed as grms. S per day.)
Intake
Outgo
Organic S .
Inorganic S
Organic S
.
Inorganic S
Increase of Organic S
Decrease of Inorganic S
Sulphur Balance
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
Without sulphate supple-
ment
G
2.772
2.212
3.127
1.837
0.355
0.375
0.020
H
2.867
2.287
3.199
1.893
0.332
0.396
0.064
With maximum sulphate
supplement
G
2.492
5.715
3.499
4.575
1.007
1.140
0.133
H
2.647
5.875
3.727
4.623
1.080
1.252
0.172
In this table due allowance has been made for sulphate in faeces. The import-
ant point to note is that the intake and outgo of sulphur balance remarkably well,
thus proving that appreciable loss in the form of H 2 S does not occur with this type
of ration.
The result has been obtained with all the precautions already noted and with
the advantage that the sulphate in the fodder was low. It is concluded from these
results that inorganic sulphate has been metabolised in some way.
234 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, III
(d) Subsidiary experiments with sheep and rabbits.
Sulphate balance experiments with sheep. The results of two tests are sum-
marised in the accompanying table :—
TABLE VIII.
Sulphate balance with sheep.
Sheep No. 1 . {
Sheep No. 2 . {
Food Consumption grms. dry matter
daily
Test
1
2
1
2
B. Hay
978
719
607
525
D. Hay
198
157
134
112
Concent.
168
176
124
125
Sulphate expressed as grms. S
daily
Ingested
0.970
0.753
0.635
0.544
Excreted
0.738
0.792
0.534
0.555
Loss
+0.232
—0.039
+0.101
—0.011
In the first test there occurred the normal loss of sulphur, which has been
observed in the bullock experiments. In the second test, a gain has been found
with both animals. This is a most interesting reversal, for which no explanation
can be given at present.
The only notable circumstance is that the animals did not thrive. They
gradually went off their food and were eating less during the 2nd test, as the state-
ment shows. It should be noted as a significant fact that both animals actually
excreted more sulphate in the 2nd test in which the intake was lower.
(e) Sulphate balance experiments with rabbits.
Two sets of determinations were made with rabbits. In the first experiment
the ration consisted of green grass, cabbage leaf and carrots. To simplify the con-
ditions, grass only was fed in the 2nd experiment. The results of the two experi-
-ents are summarised in the accompanying table :—
TABLE IX.
Sulphate balance with rabbits.
(Sulphate expressed as grms. S per day.)
Expt.
1 {
2 {
Rabbit No.
1
2
3
1
2
3
Ingested
0.2480
0.2598
0.2414
0.3337
0.3159
0.3066
Excreted
0.3341
0.3620
0.3082
0.3630
0.3498
0.3292
Gain
0.0861
0.1022
0.0668
0.0293
0.0339
0.0226
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH
CATTLE
235
The figures make it clear that, with these rations and these animals, sulphate
excretion greatly exceeds the sulphate ingested. It would appear that green
fodders contain a considerable amount of organic sulphur which is oxidised to
sulphate in the course of metabolism.
It should be noted, however, that there is a marked difference between the
results of the two tests. In the first test much less sulphate was ingested and yet
the excretion is almost as high as in the second test.
EXCRETION OF INGESTED ORGANIC SULPHUR.
From the foregoing bullock experiments the question arises whether organic
sulphur must necessarily be excreted as sulphate. Obviously if sulphate is required
for conjugation, then organic sulphur would probably be deflected into this channel
for this purpose. Therefore the first test was made with an excess of inorganic
sulphate in the ration.
Groundnut-cake which has been shown by Sullivan [1930] to contain a con-
siderable amount of cystine in the protein molecule was used as the source of organic
sulphur. The experiment was divided into four parts, the rationing being as
follows :—
1. Excess of organic sulphate plus normal allowance of groundnut-cake.
2. Excess of inorganic sulphate plus double normal allowance of cake.
3. Low organic sulphate plus double normal allowance of cake.
4. Normal allowance of cake. No sulphate.
The results of these experiments are summarised in Table X.
TABLE X.
Influence of concentrate ration on urinary sulphur.
Concentrate allowance
Sulphate in fodder
Sulphate in supplement .
.
.
.
Total sulphate in ration .
Total sulphate in urine .
.
.
.
Ethereal sulphate in urine . . .
Neutral S in urine .
.
.
.
Sulphate expressed as grms. S daily
With sulphate supplement
N
2.419
3.376
5.795
4.365
2.226
0.475
2N
2.109
3.376
5.848
4.538
2.655
0.440
No sulphate supplement
N
2.250
. .
2.250
1.717
1.717
0.131
2N
2.562
. .
2.562
1.985
1.895
0.152
236 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, III
The following points have to be noted :—
(a) Sources of phenolic bodies in urine.
The fodder consumption being reasonably constant in these tests, it is possible
to estimate roughly the phenols produced by doubling the concentrate. From
experiments 1 and 2, doubling the concentrate increased the ethereal sulphates by
.4 grms. S. As sulphate was present in excess in both these urines, this increase
of ethereal sulphate represents p. cresol together with other phenolic substances.
From experiments 3 and 4, in which sulphate was deficient (there was no free
sulphate in either of these urines), doubling the concentrate increased the ethereal
sulphate by .2 grm. S, which is attributable to p. cresol in this case. Hence the
following calculation can be made for the normal ration.
TABLE XI.
Production of phenols from hay and concentrate.
From hay plus concentrate
.
.
.
.
.
.
From concentrate . . . . . . . .
From hay . . . . . . . . .
Phenols as grms. S per day
P. cresol
1.70
0.20
1.50
Other
phenolic
bodies
0.60
0.20
0.40
Total
2.30
0.40
1.90
As the concentrate formed roughly 1/20 of the ration, clearly, it yields relatively
a much larger amount of phenolic substances. On the other hand, with the rations
actually fed, the hay provided the major part of the phenols.
(b) Source of sulphur in urine.
The first series of tests with sulphate supplement, shows that doubling the
concentrate has increased total SO 4 in urine by 0.173 grm. S, but owing to increase
of total phenols the ethereal sulphate has increased by .429 grm., which suggests
that the increase of total sulphate is due to increase of ethereal sulphate. In this
test the neutral sulphur has not been increased.
The second series of tests without sulphate supplement, shows that doubling
the concentrate has increased the total sulphate in the urine by 0.268 grm. S, but
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH
CATTLE
237
this increase is entirely in the form of ethereal sulphate due to production of more
p. cresol. The neutral sulphur is decidedly higher here with the doubled concen-
trate. This may be due in part to the fact that immediately preceding experiment
4, the animals had lived in sulphur sufficiency, while experiment 3 was carried out
after a period of sulphur deficiency. It must be admitted that the evidence from
these experiments is vitiated by the ethereal sulphates. There is an increase in
total sulphates and it is impossible to assert, though it seems likely, that this is
due mainly to the increase in ethereal sulphates. It can be said definitely, how-
ever, that the increase in total sulphates only accounts for a part of the extra
organic S (.72 grm.) in the doubled concentrate ration.
EXPERIMENT WITH REDOUBLED CONCENTRATE plus MAXIMUM SULPHATE.
Only one animal, H, was used for this test. The average results obtained from
daily determinations for 6 days were as follows :—
Sulphate in urine expressed as grms. sulphur per day.
Total Sulphate
6.060
Free Sulphate
2.591
Ethereal Sulphate
3.469
These results obtained with 4 times the normal concentrate have to be com-
pared with the corresponding data obtained with the normal and double concen-
trate allowances. The figures are as follows :—
TABLE XII.
Influence of concentrate sulphur on urinary sulphate.
—
1. Normal concentrate maximum sulphate .
2. 2 Normal concentrate maximum sulphate.
3. 4 Normal concentrate maximum sulphate
Grms. S
In concen-
trate
0.72
1.44
2.88
Increase
.72
1.44
Sulphate
in urine
(grms. S)
4.368
4.632
6.060
Increase
0.264
1.428
The noteworthy point in these results is that in experiment 2, the increase in
total urinary SO 4 is small compared with the increase of concentrate S ; while in
238 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, III.
experiment 3, the increase of total urinary sulphate approaches the increase of
concentrate sulphur. These results show that a new form of excretion has been
attained in the last test. The conclusions to be derived from the series of experi-
ments are :—
1. The sulphur of groundnut cake is digestible and metabolisable, for it has
appeared in the urine almost quantitatively in the last test.
2. In the last test there appears to be a luxury allowance of S and the luxury
allowance is completely oxidised and excreted as sulphate.
3. If fed in small amounts, groundnut cake sulphur does not appear in the
urine in any considerable proportion. Therefore, in such cases, it must be meta-
bolised in some other manner and eliminated in the faeces.
It seems to be a significant fact that the fate of the lost sulphate is similar to
that of the organic sulphur in groundnut cake when the latter is fed in small
amounts. Certainly, the sulphur from both these sources is present in the faeces,
as very carefully conducted balance experiments have shown.
Further evidence from recent experiments.
(1) Sulphate balance experiments were carried out with rice straw and sorghum
fodder, each being tested (a) without a sulphate supplement, (b) with maximum sulp-
hate supplement. Concentrate was fed throughout at the uniform rate of 2 Normal
(1.4 grms. Organic S). The results obtained are shown in the accompanying table.
TABLE XIII.
Sulphate balance experiments with rice straw and sorghum fodder.
—
Sulphate in fodder
Sulphate in supplement .
.
.
.
Total ingested .
Excreted in urine
Loss of sulphate
Sulphate in grms. S per day
Rice Straw
a
2.381
0
2.381
2.176
0.205
b
2.747
3.376
6.123
4.718
1.405
Sorghum
a
1.808
0
1.808
1.678
0.130
b
1.693
3.376
5.069
4.008
1.061
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH CATTLE
239
There is no trace of the 1.4 grms. of organic sulphur in these urines. The
usual loss of sulphate takes place when an excess is provided in the food. The
remarkably small loss in experiment (a) with sorghum is no doubt due to the fact
established by the author that sorghum produces a large excess of p. cresol, which
in this experiment has bound the sulphate effectively and thus prevented loss.
(2) Experiments with linseed cake and hay. The following series of tests were
carried out :—
(a) Normal cake no sulphate.
(b) Normal cake maximum sulphate.
(c) 2 Normal cake maximum sulphate.
(d) 4 Normal cake maximum sulphate.
The sulphate balance was determined at each stage. The results were as
follows :—
TABLE XIV.
Sulphate balances with rations of linseed cake and hay.
—
Sulphate in fodder
Sulphate in supplement
....
Total sulphate ingested .
Excreted in urine
Loss of sulphate
Sulphate in grms. S per day
a
2.650
0
2.650
1.897
0.753
b
2.586
3.376
5.962
4.320
1.642
c
2.746
3.376
6.122
4.732
1.390
d
2.613
3.376
5.989
5.404
0.585
The significant points are :—
Test (b). Normal Concentrate + Sulphate. Very high loss of sulphate. Organic
sulphur is not oxidised.
Test (c). 2 Normal Concentrate + Sulphate. Loss is slightly reduced, which
means that a small amount of organic sulphur (.25 grm.) has been oxidised.
240 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, III
Test (d). 4 Normal Concentrate + Sulphate. Loss of sulphate is very greatly
diminished. Much organic sulphur has been oxidised (1.057 grms.).
The results resemble those obtained earlier with groundnut cake. Again up
to a certain point the organic sulphur has little or no effect. Feeding in excess of
this amount causes a large increase in urinary sulphur.
SUMMARY.
1. Total sulphur and sulphate balance experiments have been carried out with
bullocks using two types of fodder.
2. The following distinctive features of the urine were noted :—
(a) Ragi straw ration. The urine contains excess of sulphate. The volatile
phenol accounts for most of the ethereal sulphate.
(b) Mature hay ration. The urine shows a deficiency of sulphate. There is
no free sulphate and the ethereal sulphate corresponds approxi-
mately to the volatile phenol. This urine contains other phenolic
bodies which conjugate with sulphuric acid if excess of the latter is
provided.
3. Numerous sulphate balance experiments with both rations invariably showed
a loss of sulphate. As far as the author is aware, this phenomenon has not been
described before.
4. Total sulphur balance experiments showed that this lost sulphate sulphur
was present, mainly in the faeces, in some other form and was not lost as H 2 S.
5. A feature of these rations is that the amount of sulphide formed in the
intestinal tract is exceedingly small. Further it has been noted that the amount
formed is related to the nature of the fodder and not to the quantity of sulphate in
the ration.
6. The amount of inorganic sulphate in the faeces is normally very small. The
amount is slightly increased when a large sulphate supplement is added to the
ration.
7. Concerning the elimination of organic sulphur (Groundnut cake) it has been
found that when small quantities are fed, very little of this S is excreted in the
urine. The major part passes out in the faeces.
On increasing the amount fed, the whole of the extra organic S is found as
sulphate in the urine.
These results show that the organic S of groundnut cake is digestible and that
there are two courses of elimination, urinary excretion as sulphate presumably re-
presenting luxury consumption.
8. Perhaps it is a significant fact that the fate of the lost sulphate is similar to
that of the organic S in groundnut cake when the latter is fed in small amounts.
SULPHUR AND SULPHATE BALANCE EXPERIMENTS WITH CATTLE
241
That the sulphur from both these sources is excreted in the faeces supports the view
that some inorganic sulphate is metabolised.
9. Subsidiary experiments with sheep and rabbits have shown that with the
former loss of sulphate can occur as with bullocks. The balance may, however,
suffer reversal.
With the rabbits and the rations employed with rabbits, sulphate excretion ex-
ceeded the intake. It was noted, however, that high excretion may occur when the
intake is considerably reduced. These observations suggest that the factors govern-
ing sulphate excretion require study.
REFERENCES.
Abderhalden and Samuel (1905). Z. Physiol. Chem. 46, 187.
Aitken (1930). Biochem. J. 24, 250.
Dennis and Leeke (1925). J. Biol. Chem. 65, 565.
Hertz, Cook and Schlessinger P. R. S. (1908). Med. London 2, 23.
Neuberg and Simon (1925). Biochem. Z. 156, 365.
Rosenfeld (1925). Biochem. Z. 157, 434.
Sullivan (1930). U. S. Public Health, R. Supplement. 66.
C