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uipped with apochromatic objectives and compensating oculars, giving magnifications of approximately 90, 180, and 500 diameters. These magnifications can be obtained by the use of 16
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he sample to stand not less than 15 minutes after mounting. Employ a magnification of about 500, which may be obtained by the use of an 8 mm. Zeiss apochromatic objective with X18 Zei
Of especial interest is the refractive constant of the filtered liquor,
shown in the last column of Table 2. The refractive constant of the
various samples is much more uniform than might be expected from a
product of this nature.
Table 2 is chiefly interesting as affording the data from which Tables
4 and 5 were calculated. The uniformity of the relations shown in
Table 5 is such that it is usually possible from one determination on
the filtrate and the determination of solids in the pulp by drying to
distinguish pulp made from whole tomatoes from that made from trimming
stock. For instance, if the specific gravity or index of refraction of
a filtrate prepared from a pulp of unknown origin, and the per cent
of solids in the pulp by drying, do not agree approximately with the
relation between these determinations as shown in Table 5, it may be
assumed that the sample was not prepared from whole tomatoes, or that
some other substance, such as salt, has been added. Moreover, trimming
stock pulp rarely conforms to the relations found in whole tomato pulp.
For instance, the insoluble solids are usually higher and the acid
lower in trimming stock pulp.
Trimming Stock Pulp
In Table 3 are given the results of the examination of 21 typical
samples of trimming stock pulp prepared at different plants and in
different localities. This table is of especial interest in showing
that the relations between the results of the various analytical
determinations differ from those of whole tomato pulps as given in
Table 5. For instance, in No. 1470 the immersion refractometer reading
is 45.90, and the per cent of solids is 9.54, whereas, according to
Table 5, the per cent of solids in the pulp corresponding to an index
of refraction of 45.90 should be 8.57. The specific gravity of the
pulp is 1.0373, which, according to Table 5, should correspond to 8.98
instead of 9.54. Of course it cannot be said definitely that a pulp
which on examination is found to conform to all the relations shown
in Table 5 is necessarily whole tomato pulp. It is entirely possible
for an occasional sample of trimming stock pulp to conform to all the
relations shown in that table; moreover, the extent to which different
samples of trimming stock pulp will vary from the relations shown in
Table 5 differs with the manner of preparation. For instance, if a
portion of the juice is discarded in the manufacture of trimming stock
pulp, as is still the practice of some manufacturers, the variation
from whole tomato pulp will be greater than otherwise and the variation
will increase with the amount of juice discarded.
Methods of Analysis[2]
These methods may also be applied to the examination of raw tomatoes
and canned tomatoes. In applying the relations given below to the
results obtained by the examination of tomato pulp or canned tomatoes,
it is assumed that no substance such as sugar or salt has been added.
If salt is found to be present in excess of the amount normal to
tomatoes (from 0.05 to 0.1 per cent), it is necessary to determine the
amount and make correction therefor before applying the relations given
below.
[2] This chapter is largely taken from an article by Bigelow and
Fitzgerald, published in the Journal of Industrial and Engineering
Chemistry. The section on Microscopic Examination is substantially
a reprint of an article by Bigelow and Donk published in the trade
papers in September, 1918. For further information on the analysis of
tomato products the Methods of Analysis of the Association of Official
Agricultural Chemists (1919) should be consulted.
In examining raw tomatoes, care must be taken to secure a
representative sample of the juice. This cannot be done by applying
pressure directly, as the juice of the seed receptacles is of different
composition from that of the fleshy part of the tomato. It is
necessary, therefore, to crush the sample and thoroughly cook it in a
flask surrounded by boiling water and connected with a reflux condenser.
Microscopic Examination
The laboratory of the National Canners’ Association is frequently asked
to examine samples of tomato products to determine whether or not they
comply with the Government requirements. In examining these samples we
use the Government method (the Howard method), but do not participate
in the discussions regarding its merits and shortcomings.
It is our experience that skilled analysts can check themselves and
each other with reasonable accuracy, and it is our duty to tell the
manufacturer whether his product is legal. Should the Bureau of
Chemistry adopt some other method as preferable to the Howard method,
it would be our duty to use the new method and continue to serve the
industry by telling the manufacturer whether samples submitted by him
would pass the Government tests.
With a full understanding of our attitude in this matter many
manufacturers of tomato products send samples from time to time for
examination. It is made plain in every instance that the results
obtained by the examination of a particular sample refer only to the
batch from which that sample was taken and may give no indication of
the character of any other batch.
Some manufacturers of tomato products use the Howard method as a check
on their factory control. For this purpose it is not satisfactory to
have samples examined in a laboratory located at a distance from the
factory. Even if several samples are examined from a day’s run, they
probably do not represent all the pulp manufactured on that day. It
sometimes happens that one wagonload of tomatoes is almost entirely
free of rotting material, whereas the succeeding load contains a
considerable amount. Even with inefficient sorting, the pulp made
from the first load will show a low microscopic count whereas,
unless sorting is exceptionally good, the pulp made from the second
load may show a high count. Thus one batch may readily comply with
the requirements of the Bureau of Chemistry and the next batch may
be outside of those limits. Because of this fact this laboratory
recommends that manufacturers of tomato pulp do not rely upon the
microscopic results of a single sample. The only way in which the
product may be absolutely controlled by means of the microscopic count
is to examine a sample from each batch—that is, from each kettleful or
tankful that is evaporated. This is manifestly impossible. It would
require several analysts for one plant. Moreover, it is entirely
unnecessary.
It has been found that much better results can be secured by having
an analyst in the plant to examine samples from time to time. Then,
whenever the microscopic count becomes excessive, he can locate the
trouble and see that it is corrected.
Manufacturers who desire frequent analyses of their products,
therefore, should employ an analyst and arrange to have him instructed
in a laboratory conversant with the Howard method as used by the
Government. The laboratory of the National Canners’ Association makes
it a practice to give the necessary instruction in this method to
analysts employed by members of the association. These analysts should
be carefully selected. Other things being equal, better results should
be expected of a college graduate or at least one who has had college
training in biology and chemistry. It has been repeatedly demonstrated,
however, that a carefully selected man or woman with common school
education can learn the method and use it with sufficient accuracy for
factory control. The person selected for this work should have good
powers of observation and a positive character.
This laboratory has heretofore advised that manufacturers of tomato
pulp should not give too much attention to the microscopic count of
their product. We have maintained that the expense would be better
placed on the sorting belt; that if the sorting and trimming were
adequately done, the plant maintained in a sanitary condition and the
product manufactured as rapidly as possible, a low microscopic count
would be assured. This we still maintain is true. So many cases have
come to our attention, however, in which canners have not succeeded
in maintaining the degree of sorting necessary with a product of this
kind that we have grown to feel that the presence of an analyst working
continuously in a plant is an additional safeguard.
The conditions attending the canning of tomatoes are widely different
from those attending the manufacture of tomato pulp. The ordinary rot
is almost always apparent from the outside of the tomatoes[3] and is
removed by the peelers when preparing tomatoes for canning. Practically
none of it, therefore, finds its way into the can. With pulp it is
quite different. Any rot which is not removed by sorting and trimming
goes into the cyclone and passes into the pulp. With trimming stock
pulp, the condition is obviously much worse than with whole tomato
pulp. One hundred pounds of tomatoes will yield not far from 85 pounds
of cyclone juice. If only trimming stock is made into pulp, however,
nearly half the tomatoes are used for canning and the remainder (50 or
55 pounds of trimming stock) will only make something like 35 or 40
pounds of cyclone juice. Yet, since the rot is almost entirely on the
outside of the tomatoes, this 35 or 40 pounds made from the trimming
stock contains the same amount of molds as the 85 pounds manufactured
from the whole tomatoes. The mold count of the trimming stock pulp,
therefore, is much higher than that of whole tomato pulp made from the
same raw product.
[3] Tomatoes are sometimes found with rotten centers of which there
is little or no external evidence. This is unusual, however, and
the influence of this form of rot under manufacturing conditions is
negligible.
The Bureau of Chemistry condemns tomato pulp whose microscopic
examination gives results as high as the following figures:
Molds 66 per cent of fields.
Bacteria 100 million per cubic centimeter.
Yeasts and spores 125 per 1/60 cubic millimeter.
These figures, of course, apply to the Howard method as employed by
the Bureau of Chemistry. The method is entirely arbitrary and results
agreeing with those obtained by the Bureau of Chemistry can be obtained
only by using this method substantially as it is used by the bureau.
An examination of the pulp, therefore, by an analyst who is not
thoroughly conversant with this method as it is employed by the Bureau
of Chemistry not only is useless but may actually afford a manufacturer
a false sense of security which will be greatly to his disadvantage.
Microscopic Equipment Required
The apparatus employed by the Bureau of Chemistry includes apochromatic
objectives and compensating oculars. In 1914 it became impossible to
obtain these accessories[4] because of the European war and equivalent
apparatus of American manufacture was found to give the same results.
Both of these forms of apparatus are recognized in the official Howard
method which is given below.
[4] Apochromatic objectives may now be obtained but are more expensive
than the achromatic.
This laboratory made a careful study of the accessories available in
order to determine what could best be used. It was found that very
satisfactory results could be obtained by employing a 10X Huyghenian
ocular and a 4 mm. achromatic objective (working distance 0.6 mm.) and
a 16 mm. achromatic objective. These accessories require a careful
adjustment of light, but with proper use enable an analyst to secure
satisfactory results. It is found that the best results are obtained
with a rather dark field.
The apparatus necessary for the Howard method, including the
accessories mentioned above, may be obtained of two American
manufacturers, the Bausch & Lomb Optical Company, of Rochester, N. Y.,
and the Spencer Lens Company, of Buffalo, N. Y.
There is given below a full list of the optical apparatus required,
including catalog numbers of the two manufacturers, as far as numbers
have been assigned by them to the various items. In addition to the
apparatus given in this list, the analyst should have a 50 c. c.
graduated cylinder for measuring and diluting samples. This may be
obtained of any dealer in chemical apparatus and at many drug stores.
When ordering the optical apparatus the full description as given below
should be included.
Optical Apparatus for the Howard Method
Quantity Item Bausch &
desired Lomb Spencer
1 Microscope without oculars,
objectives or other a FF 44
1 Abbe condenser with two iris diaphragms
(lower and upper) 1740 300
1 Double nosepiece 1844 450
1 16 mm. achromatic objective 1021 108
1 4 mm. achromatic objective with working
distance of 0.6 mm. 1029 116
1 8 mm. achromatic objective with working
distance of 1.6 mm. 1027 112
1 10X Huyghenian ocular 1104 142
1 Mechanical stage 2116 485
1 Substage lamp with Daylite glass 1774 385-B
1 Blood counting chamber (Haemacytometer with
ruling of Thoma, Neubauer, Jappert, Brewer
or Turk) 3550 1472
6 Cover glasses for same, 20×21 mm., 0.4 thick 3595 1460
1 Howard’s mold counting chamber (with ¾ inch
inner disk) for same 3566 Special
6 Cover glasses for same 33 mm. square,
0.6 mm. thick 3598 Special
2 Cases for counting chambers 3580 1505
All analysts undertaking the Howard method should secure copies of the
two bulletins of the United States Department of Agriculture written
by Mr. B. J. Howard—Bulletin 569 on Sanitary Control of Tomato Canning
Factories and Bulletin 581, Microscopic Studies on Tomato Products.
These bulletins may be obtained from the Superintendent of Documents,
Government Printing Office, Washington, D. C., on payment of five cents
each in coin.
The details of the method as given below are reprinted from the Methods
of Analysis of the Official Agricultural Chemists as amended in 1921.
Apparatus
(_a_) _Compound microscope_.—Equipped with apochromatic objectives and
compensating oculars, giving magnifications of approximately 90, 180,
and 500 diameters. These magnifications can be obtained by the use
of 16 and 8 mm. Zeiss apochromatic objectives with X6 and X18 Zeiss
compensating oculars, or their equivalents, such as the Spencer 16 and
8 mm. apochromatic objectives[5] with Spencer X10 and X20 compensating
oculars, the draw-tube of the microscope being adjusted as directed
below.
(_b_) Thoma-Zeiss blood counting cell.[2]
(_c_) Howard mold counting cell.—Constructed like a blood-counting
cell but with the inner disk (which need not be ruled) about 19 mm. in
diameter.[6]
[5] We are informed that the Bausch and Lomb Optical Co. also furnishes
suitable apochromatic objectives and compensating oculars for use in
counting molds, yeast and bacteria by the Howard method.
[6] Comment by authors: In using these cells the plane parallel cover
glasses furnished with them by maker should be used instead of the
ordinary microscope cover-glasses, since the latter are subject to
curvatures that introduce errors in the thickness of the mounts.
Molds.—Tentative
Clean the special Howard cell so that Newton’s rings are produced
between the slide and the cover-glass. Remove the cover and place, by
means of a knife blade or scalpel, a small drop of the sample upon the
central disk; spread the drop evenly over the disk and cover with the
cover-glass so as to give an even spread to the material. It is of the
utmost importance that the drop be mixed thoroughly and spread evenly;
otherwise the insoluble matter, and consequently the molds, are most
abundant at the center of the drop. Squeezing out of the more liquid
portions around the margin must be avoided. In a satisfactory mount
Newton’s rings should be apparent when finally mounted and none of
the liquid should be drawn across the moat and under the cover-glass.
Place the slide under the microscope and examine with a magnification
of about 90 diameters and with such adjustment that each field of
view covers 1.5 sq. mm. This area is of vital importance and may be
obtained by adjusting the draw-tube in such a way that the diameter of
the field becomes 1.382 mm. as determined by measurement with a stage
micrometer.[7] A 16 mm. Zeiss apochromatic objective with a Zeiss X6
compensating ocular or a Spencer 16 mm. apochromatic objective with a
Spencer X10 compensating ocular, or their equivalents, shall be used
to obtain this magnification. Under these conditions the amount of
liquid examined is .15 cmm. per field. Observe each field as to the
presence or absence of mold filaments and note the result as positive
or negative. Examine at least 50 fields, prepared from two or more
mounts. No field should be considered positive unless the aggregate
length of the filaments present exceeds approximately one-sixth of the
diameter of the field. Calculate the proportion of positive fields
from the results of the examination of all the observed fields and
report as percentage of fields containing mold filaments.
[7] Comment by authors: Obviously after the proper draw-tube length has
been secured that adjustment should be noted and always used in making
mold counts.
Yeasts and Spores.—Tentative
Fill a graduated cylinder with water to the 20 cc. mark, and then add
the sample till the level of the mixture reaches the 30 cc. mark.
Close the graduate, or pour the contents into an Erlenmeyer flask,
and shake the mixture vigorously for 15 to 20 seconds. To facilitate
thorough mixing the mixture should not fill more than three-fourths of
the container in which the shaking is performed. For tomato sauce or
pastes, or products running very high in the number of organisms, or
of heavy consistency, 80 cc. of water should be used with 10 cc. or 10
grams of the sample. In the case of exceptionally thick or dry pastes,
it may be necessary to make an even greater dilution.
Pour the mixture into a beaker. Thoroughly clean the Thoma-Zeiss
counting cell so as to give good Newton’s rings. Stir thoroughly
the contents of the beaker with a scalpel or knife blade, and then,
after allowing to stand 3 to 5 seconds, remove a small drop and place
upon the central disk of the Thoma-Zeiss counting cell and cover
immediately with the cover-glass, observing the same precautions in
mounting the sample as given under 28.[8] Allow the slide to stand
not less than 10 minutes before beginning to make the count. Make the
count with a magnification of about 180 diameters to obtain which the
following combination, or their equivalents, should be employed: 8 mm.
Zeiss apochromatic objective with X6 Zeiss compensating ocular, or an
8 mm. Spencer apochromatic objective with X10 Spencer compensating
ocular with draw-tube not extended.
Count the number of yeasts and spores[9] on one-half of the ruled
squares on the disk (this amounts to counting the number in 8 of the
blocks, each of which contains 25 of the small ruled squares). The
total number thus obtained equals the number of organisms in 1/60,000
cc. if a dilution of 1 part of the sample with 2 parts of water is
used. If a dilution of 1 part of the sample with 8 parts of water is
used the number must be multiplied by 3. In making the counts, the
analyst should avoid counting an organism twice when it rests on a
boundary line between two adjacent squares.
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