FERROMAGNETIC MICROTRACERS AND THEIR USE IN FEED
APPLICATIONS
Nikolay Barashkov, David Eisenberg, Sylvan Eisenberg, Joseph Mohnke
Micro Tracers., Inc, 1370 Van Dyke Ave., San Francisco, CA 94124, United States
ABSTRACT
This presentation is devoted to a wide variety of analytical ferromagnetic tracers used to assure the quality
of mixed formula animal and poultry feeds. When formulated in vitamin, mineral or medicated premixes, a
MicrotracerTMr serves to mark the presence of the premix in the finished feeds as well as to identify the
premix and feeds containing it as proprietary. When assayed quantitatively, a MicrotracersTM can be used to
document efficacy of mixing as well as adequacy of batch to batch “cleanout” of mixers and other feed
manufacturing equipment.
Iron-based tracers coated with food dye(s), as well as the recent development of several new types of
ferromagnetic microtracers will be discussed. One new type includes multicolored dots on the surface of
ferromagnetic particles. These tracers can be included in animal feed and premixes, granular products and
bulk materials, then retrieved by means of the Rotary DetectorTM - laboratory magnetic separator and then
identified with a microscope or other magnifying device. Another recently developed and patented tracer
consists of printed numerical and letter code information (size of letters about 25-50 microns) on iron
particles. This technology employs a new lithography process only using ingredients approved by FDA for
human and animal consumption. Easily retrievable and recognizable with a microscope, these new tracers
can be used to code dry mixes as proprietary.
Another new tracer consists of iron oxide - based nanoparticles containing up to 5% of cobalt oxide. They
have been quantitatively retrieved from liquid feeds in lab and industrial trials. Nanoparticles have been
used for making liquid ferromagnetic tracer according to procedure that includes the synthesis of
ferromagnetic liquid, separation of this liquid from organic phase, dispersion of ferromagnetic liquid in
aqueous solution of surfactant and determination of cobalt content in final product. New tracer can be used
to estimate the concentration of enzymes in a liquid feed, as well as for coding liquid enzymes and, in
general, for validating mixing of liquid feeds.
Key words: microtracers, ferromagnetic, iron particles, animal feed, food grade color, nanoparticles,
liquid feed
INTRODUCTION
The worldwide formula feed industry manufactures more than 400 million tons annually. Manufacturers
waste labor, energy and capital when they mix feeds longer than necessary to achieve a complete blend.
Excess mixing may also cause degradation of vitamins and medications. If feed is not completely mixed,
portions of the feed will contain either too much or too little of the formulated ingredients. This excess
variability causes economic losses to users of the feed and may increase the incidence of illegal drug
residues. Periodic routine mixer testing is both economically and ethically justified. Starting in the mid1990s regulatory authorities in many countries have become more concerned in assuring medicated feeds
are mixed completely and that all micro ingredients are added as formulated.
Failure to consistently add the formulated levels of micro ingredients may be a greater problem than mixing
in technologically advanced countries, where computerized micro-bin systems are commonly used to add
micro ingredients. Feed manufacturers owe a response to the public to ensure their feeds are properly
manufactured. This interest coincides with their self-interest to make the best product reasonably possible.
This benefits the feed manufacturer's customers and the feed manufacturer himself if he is an integrated
producer. By developing and implementing well-designed sampling and analysis plans the feed
manufacturer can validate his mixing and micro ingredient addition operations.
Different substances including amino acids, microminerals [1], salt [2], and drugs [3] have been used as
tracers to measure feed mixing efficiency. More exotic tracers for livestock feeds include chromium
mordanted fiber for dry feeds and cobalt-ethylene diaminetetracetic acid for liquid feeds [4].
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For many years, the colored iron tracer particles which can be separated from the formula feeds
magnetically have been successfully used in practice. Patents assigned to Micro Tracers, Inc. [5-9] disclose
the composition of tracers consisting of iron or other ferromagnetic material coated with certified FD&C
dyes. These tracer particles can be counted. Even major losses of the food color due to abrasion or diffusion
can be tolerated without loss of accuracy in the particle count as 20% of the formulated color will still yield
a countable spot. In animal feed applications, essential nutrients or drugs may themselves be used as
identifiable coatings. These tracers can be separated from a mix much faster and more clearly than prior
tracers isolated by sedimentation in perchloroethylene. The particulate ferromagnetic material passes a U.S.
40 mesh screen but is retained by a U.S. 325 mesh screen. Finer or coarser ranges may be preferred
depending upon the nature of the dry mix to be marked with the tracer.
Coated tracers may be produced by first dry-blending water soluble FD&C colors with the iron grit carrier.
Water is then added to dissolve the dye. The mix is then dried, and screened. The product is then analyzed
for conformance to specification then packaged as finished product. The screening may be varied
depending upon the application. Certified FD&C dyes such as Blue #1, Blue#2, Red #2, Red #3, Red #40,
Yellow #5, Yellow #6 and others serve as easily identifiable coating materials. Mixtures of dyes differing
in diffusion rates may be used to increase the number of different tracers and to produce truly unique
markers. For example mixtures of Blue #1 and Yellow #6 produce blue streaks or rings with yellow nuclei
by the detection procedure described herein below. Mixtures of Blue #1 and Red #2 produce blue streaks or
rings with red nuclei.
Iron-based MicrotracersTM include the following: 1. Microtracer F (Colored Iron grit, 25,000 particles per
gram); 2. Microtracer FS (Colored Stainless steel grit, 50,000 particles per gram); 3. Microtracer RF
(Colored Reduced iron powder > 1,000 000 particles per gram). These tracers may be detected and
quantified using a Rotary Detector - laboratory magnetic separator or by use of special magnetic probe.
Below we will consider two applications of iron-based tracers coated with single color food grade dye
which include the validating the mixing process and testing for "Cross Contamination" in Medicated Feeds.
Besides traditional tracers, the recent development of several types of new types of microtracers will be
discussed.
RESULTS AND DISCUSSION
1. Validating the mixing process.
At least five issues must be considered in validating the mixing process [10,11]: 1. Selection of - one or
more tracers; 2· Addition of the tracer to the test feed; 3· Sampling the feed; 4· Analysis the samples; 5·
Interpreting the results.
I.1. Selection of the tracer. Whatever analyte is chosen for the mixer test, it will then be used as a tracer
for all other ingredients. If it yields results typical of a complete mix, one will assume all other ingredients
are mixed completely. This may not always be a correct assumption. Powdered feed additives may become
electrostatically charged and stick to the walls of mixers that accumulate as clumps that drop off rarely. If
one tests for this ingredient as a tracer, analytical results will be consistent among samples but always low
on average unless one happens to test the rare clump. Such an analyte would not accurately reflect the
mixing of other ingredients and would probably be an inappropriate choice for use in evaluating mixer
performance.
At least the following criteria should be considered in selecting the tracer: A. The tracer should be
contributed from only one source; B. The tracer should be a microingredient; C. There should be an
analytical procedure to determine the tracer of known or determinable accuracy and precision; D. The
analytical procedure should be inexpensive; E. The analytical procedure should be quick: ideally one that
may be performed "on-the spot"; F. One should be able to interpret results objectively.
Vitamin and drugs assays may be used to validate completeness of mix, but they are
expensive, are often subject to considerable analytical error and often require weeks
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before analytical results are reported. Salt is often used as a tracer to evaluate mixing by determining either
sodium or chlorides in feeds. Salt, however, has significant deficiencies as a tracer. It is added at five to 20
kilos per metric tonne and is there by hardly a microingredient. It may not be reasonable to assume mixing
of a medication added at three parts per million is correct based upon acceptable results for salt added at
20,000 parts per million. Further, sodium and chloride may be contributed to feeds by other ingredients,
yielding background "noise" confusing interpretation of results.
Minerals and amino acids are also widely used and these largely meet the criteria outlined in this paper and
generally yield meaningful information. The cost of analysis for minerals such as manganese and zinc by
atomic absorption spectrophotometry is often low, possibly $20/sample if preformed by a commercial
laboratory or less if performed by the feed manufacturer. Reproducibility of results on a given sample is
often good with a coefficient of variation of five to eight per cent possible. Some minerals are contributed
to feeds in significant quantities from only one source: the mineral premix. Amino acid analyses by HPLC
may be reproducible with a coefficient of variation of five per cent or less, although they may be more
costly to perform than mineral analyses. MicrotracersTM F are also used widely to evaluate mixing. These
non-nutrient particulate tracers are designed to satisfy the criteria outlined in this paper. The analytical error
in their determination may be two to three per cent and they may be performed "'on-the spot" by
technicians with comparatively little training.
I.2. Addition of the tracer to the test feed. This should be via a premix possibly made by mixing the
tracer by hand with other common feed ingredient. The amount of premix added to the test feed should be
similar to the lowest addition ingredient normally formulated in the feed. If medicated premixes added at
500 grams/ tonne then the tracer premix might be added at this level. The location of tracer addition is
usually where other microingredients are added. Test results will then validate existing procedure, the
tracer may also be added at other locations to determine if the location of microingredient addition has a
significant effect on the time required to achieve a complete mix. In several tests, it took 30 second or less
to achieve a complete mix when a tracer was added into the center of the mixer is compared with when it
added at the end of the mixer.
1.3. Sampling the feed. In validating mixer performance "grab" samples should ideally be taken from
within the mixer. The samples should be carefully identified and all test parameters should be carefully
recorded. Samples should be adequate in size to permit repeat analysis samples evidencing unusual results.
How many samples should be taken? Taking one sample is an infinite improvement over taking none. We
usually take ten samples from each of five consecutive batches and believe this adequate to detect major
deviations from complete mixing that could lead to significant economic losses or regulatory compliance
problems.
1.4. Analysing the samples. One should take a given weight of sub-sample from each sample for analysis
without homogenizing the sample. This makes the "level of scrutiny" of the test the weight of the subsample analyzed rather than the weight of the sample taken. It makes the test more severe, increasing the
likelihood of a filling conclusion but also improving the likelihood a favorable result is correct. If mixing is
good the worst of conditions, then it should be good under less severe ones. The ideal "level of scrutiny"
would be batches of feed consumed by target animals poultry or fish at one feeding.. The ideal "level of
scrutiny" for the tracer would be one added at the lowest level of any microingredient formulated in the
feed, though this issue is complex and effected by intermediate mixing procedures utilized to achieve
adequate dispersion. For example, critical microingredients may be dissolved in a liquid and sprayed onto a
dry carrier to facilitate achieving adequate dispersion. Samples should then be analyzed using appropriate
methodology, proper instrumentation and skilled personnel.
1.5. Interpreting results. This should be done by comparing the coefficient of variation found from the
test data with the coefficient of variation inherent in the method. The method coefficient of variation is
what one would expect from repeat analysis of the same sample. Using a maximum permitted coefficient of
variation of 10 per cent is arbitrary and capricious unless it can be related to the variability of the analysis.
If a coefficient of variation of two per cent can be achieved from a method, this could be used as the goal
for judging a mix complete. If a coefficient of variation of 15 per cent was the best that could be achieved
3
front a method, this could be used as a goal for judging a mix complete. Such standards would be
unrealistically high however, as 50 per cent of all tests of a complete mix would evidence more variation
than the goal. A more realistic goal would be to consider results acceptable if they would occur by chance
from a complete mix in at least one per cent of all tests of such a mix. This may mean considering data as
evidencing a compete mix when it yields a coefficient of variation is 50 per cent or more greater than the
method coefficient of variation. In evaluating particulate tracer counts, one may utilize Poisson statistics to
determine if mixing is complete. If one makes no allowance for analytical error, if a mix is complete it
should yield test results with I standard deviation equal to the square root of the average count. An average
count of 100 should yield a standard deviation of 10 and a coefficient of -variation of 10 per cent. An
average count of nine would yield an expected standard deviation of three and an expected coefficient of
variation of 33 per cent.
2. Testing for "Cross Contamination" in Medicated Feeds.
Medicated animal, poultry and aquatic feeds are often manufactured at feedmills making feeds for many
species or if not at least many formulations for one species. Contamination of medications to non-target
feeds is inevitable but quantifiable and controllable. Medicated feed assays, unfortunately, often do not
offer a viable control mechanism because: they are expensive, they cannot be performed immediately and
they are often inaccurate at contamination levels (i.e. 1 % or less of formulated levels) [12].
Microtracers F offer a vehicle for locating and quantifying "cross contamination" of medicated feeds.
Instead of running expensive drug assays, one uses one or more Microtracers F as indicators for the coded
medication. The tracer is first mixed into the medicated feed premix, then the premix is added to the feed
and mixed. Many samples are taken either only at truck loading or at many locations, depending upon the
scope of the study. It is critical that feeds not formulated with the microtracer follow the identical route at
the feedmill as the "target" batch containing the microtracer. Samples yielding significant levels of the
tracer may then be analyzed for the medication to confirm the preliminary results provided by the tracer.
2.1. Addition of the Tracer. The microtracer is generally added to the medicated premix to yield 50
grams of tracer per metric tonne of final feed. Since the tracer has an expected count of 25,000
particles/gram, this means 50 X 25,000 or 1,250,000 colored iron particles are added to a one metric ton
batch with 1,250 particles expected per kilogram of feed or 125 per 100 grams, assuming the feed is
completely mixed, the tracer is stable and it meets its specified count.
2.2. Sampling. One should take a minimum of four and preferably ten or more "grab" samples from the
target batch formulated with the Microtracer. Samples of the target batch should weigh at least 200 grams
and ideally be taken at several points in the feed production system. If one analyzes 80 gram "grab" subsamples of each target sample, one will expect to find 100 tracer particles per sample. One should then take
samples from one or more following batches not formulated with the medication or the tracer. These
samples should weigh at least 1-kilo each. If one analyzes 800 grams of a feed supposed to contain no
tracer and finds 100 tracer particles, then if total tracer recovery is 100% of theory one may estimate 10%
of the total tracer (and thereby the drug) was found as contamination in that sample. By analyzing very
large samples of feeds supposed to contain no tracer, the "sensitivity" (ability of the tracer to allow accurate
detection of very low level contamination) may be improved by a factor of 10 or more. Individual batches
of feed must be isolated and their flow controlled. All samples must be carefully identified and detailed
records of all aspects of the test should be made.
The basic flow of feed at a feedmill includes the following: 1. feed ingredients after being ground to mash
with a hammer mill (if necessary) are gravity fed into a mixer and mixed; 2. the mixed feed is then dumped
into a surge bin. 3. the feed is then discharged from the surge bin using a screw conveyer; 4. the feed is then
elevated in the feedmill using a bucket elevator system; 5. the feed is then gravity dumped into a holding
bin over a pellet mill; 6. the feed is then pelleted and cooled; 7. the feed is then elevated again using a
bucket elevator or pneumatic system; 8. the feed is then distributed to holding bins; 9. the feed is then
loaded into trucks.; 10. the feed is then carried to farms where it is discharged into bins; 11. the feed is then
distributed from the on-farm bins to the animals, poultry or fish for consumption.
Medications are usually added as a microingredient into the mixer, sometimes by hand but often utilizing
4
computer controlled micro-bin systems. "Cross Contamination" of the medication may occur in the microbin system even before the medication reaches the mixer. "Cross contamination" to prior batches may also
occur, as when mixer discharge gates leak and an earlier batch remains in the surge bin. As an initial
investigation, one may take samples of the "non-target" following feed only at truck loadout taking care to
be certain the feed takes the identical route as the medicated feed with Microtracer. If one finds very little
or no tracer from the loadout samples, this indicates there may be very little "cross contamination"
occurring anywhere in the system. If one finds Microtracers in the "non-target" following feed, however,
much more extensive testing is required to diagnose and then have a basis for correcting the problem.
Portions of the medication may be left as "contamination" to non-target batches at all production points. To
reliably diagnose the potential for "cross contamination" of medications to non-target batches of feed, it is
critical to take multiple "grab" samples at multiple locations of both the target batch (with medication) as
well as one or more following batches. It must be re-emphasized that it is critical the following batches take
the identical route through the feedmill as the initial medicated batch formulated with Microtracer.
3. Non-traditional iron-based microtracers
Besides traditional iron-based tracers coated with single color food grade dye, several new types of
microtracers are recently developed [13-15].
3.1. Microtracers with multicolored dots. This type of microtracers includes the creation of multicolored
dots on the surface of ferromagnetic particles. These tracers were capable to withstand the conditions of
making pelleted feed, as well as manufacturing conditions applicable to meat and bone meal (pressure
cooking at 2 atmospheres and 133oC for 20 min). The microtracers comprise a finely divided iron
approximately in range from 50 to 200 microns at the broadest dimension. Below 50 microns, the code may
be difficult to read, and more sophisticated magnification apparatus may be required. According to recently
submitted patent application [13] two types of multicolored ferromagnetic microparticles are proposed.
First type of microparticles has a multicolored code information in a form of two or more colored spots and
consist of the following: a) a core comprising a ferromagnetic material, b) a masking layer of white
reflecting pigment enveloping the ferromagnetic core c) two or more coloring agents present on the
masking layer. Second type of microparticles has a multicolored code information in a form of layers with
two or more colors and it is prepared by spin coating or dip coating. The process of coating includes the
usage of the FDA approved ingredients only. Fig.1A shows the microphotograph of iron particles (40/+100 mesh) coated with a white layer of titanium dioxide and colored with two types of alumina
particles (-230/+325 mesh) colored with FD&C dyes Red#40 lake and Blue #1 Lake.
A
B
Fig.1 Microphotograph of iron particles [13,14] : A. Particles are coated with a white layer of titanium
dioxide and colored with two colored types of alumina particles; B. Particles with the image of letters (50
micron tall) printed by photo-resist process.
The new microtracers can be applied to animal feed and premixes, granular products and bulk materials,
then collected by means of the Rotary Detector TM and then identified with a microscope or other
magnifying device. Each microparticle can include at least two distinguishable colors or a plurality of
colors. If single microparticles are made using seven different colors, 2 7 or 128 binary numbers can be
created. If the same seven colors are used to make dual-color microparticles (i.e., particles having two color
layers each), 21 unique dual-color microparticles can be created (Table 1).
5
Table 1. Number of combinations from dual-color microparticles and FDA&C dyes which can be used in
order to create the corresponding coatings
##
1
2
3
4
5
6
7
Color, Dye, maximum of
absorbance
Blue , FDA&C Blue#1,
630 nm
Brown, FDA&C Brown
HT, 458 nm
Green, FDA&C Sodium
Copper Chlorophylin,
406 nm
Orange, FDA&C
Yellow #6, 482 nm
Red, FDA&C Red #40,
510 nm
Violet, FDA&C Dark
Violet, 570 nm
Yellow, FDA&C, Yellow
#5, 425 nm
Yellow
Violet
Red
Color
Orange
Green
Brown
Total
x
x
x
x
x
x
6
x
x
x
x
x
x
x
x
x
x
x
x
x
x
x
5
4
3
2
1
0
Total: 21
3.2. Microtracers with numerical and letter code information. Original procedure for printing numerical
and letter code information (size of letters about 25-50 microns) on the iron particles (Fig.1 B) has been
patented recently [14]. Proposed in this technology new photo-resist process includes the ingredients
approved by FDA for usage for human consumption and for the animal feed industry only. It was shown
that the printing process on iron particles can be accomplished with both a "positive" and a "negative"
photo-resist. In a positive working resist the monomer or oligomer is deposited on the metal surface in the
form of a viscous liquid. It is then irradiated through a mask (this is a drawing or printing on a transparent
sheet) and polymerization (crosslinking) takes place only at the exposed places. The unirradiated liquid
monomer is then washed away in a suitable solvent, and the exposed metal can be dissolved (completely or
partially) in an etching bath. Finally the protective polymer layer is removed by chemical or mechanical
means, and the printed image is ready. In a negative working resist the monomer is first polymerized over
the entire metal surface, then the protective polymer layer is irradiated through the mask. In the irradiated
areas the polymer is degradated into smaller units and becomes soluble; it can then be removed by
treatment with a suitable solvent and the etching bath will attack this exposed metal. A variation of this
technique can be used for a preparation of integrated circuits or other printed images with a resolution of
the order of one micron. In this case an exposure to light is done through a mask with an optical reduction
of the size of the image.
Easily retrievable and recognizable with a microscope, new tracer with numerical and letter code
information can be potentially used to authenticate materials for a number of different applications,
including marking animal feed and premixes, other types of particulate, granular products and bulk
materials.
3.3. Liquid tracer with ferromagnetic nanoparticles. Iron-based microtracers usually are not applicable
for validating mixing of liquid feeds, as well as for coding liquid additives, such as enzymes, and
evaluation of their distribution in premixes and final feeds. In order to meet the growing demand we have
developed and tested a magnetically retrievable liquid microtracer containing iron oxide - based
nanoparticles [15]. Nanoparticles combining iron oxide (higher 95%) and cobalt oxide (below 5%) have
been used for making liquid tracer according to a procedure that includes the synthesis of ferromagnetic
liquid, separation this liquid from the organic phase, dispersion of ferromagnetic liquid in aqueous solution
of surfactant and analytical determination of cobalt content in final product. The laboratory trial with an
addition of liquid tracer (mixable with liquid enzyme added at concentration 110 ppm to the feed) at level
100 ppm to the dry feed, followed by retrieving the ferromagnetic nanoparticles and analyzing the content
6
of cobalt oxide in them yields approximately 75% tracer recovery. The first industrial trial with the new
liquid tracer has been performed recently by our colleagues from Micro Tracers Services Europe with a
major European pig feed manufacturer. Table 2 shows the results of this trial where the relatively large
amount of liquid tracer (1 kg) has been used for evaluation of mixing quality for several liquid ingredients
in 510 kg of liquid pig feed mix. The data presented in Table 2 was interpreted using Poisson and Chi
squared statistics. Treating a series of counts (absorbance values) as a Poisson distribution, the mix is
judged complete. If the counts would occur by chance from a “perfect” mix in fewer than 5 of 100 studies,
the mix is judged “probably incomplete”. If the counts would occur by chance from a “perfect” mix in
fewer than one of 100 studies, the mix is judged incomplete.
Table 2. Results of homogeneity industrial trial with liquid microtracer (tracer recovery is about 45%)
## analyzed samples
Parameter
Concentration of
cobalt in sample
(absorbance units
at 667 nm) x 104
Mean value of
concentration
Standard
Deviation, +/Coefficient of
variation, %, +/Chi square
Probability, %
1
90
2
83
3
74
4
80
5
92
6
91
7
96
8
100
9
83
10
104
11
95
12
60
87.33
12.18
13.94
18.67
4.46
Acknowledgement. The authors are thankful to Dr. Sabine Artelt ( Micro Tracers Services Europe GmbH)
for participation in the liquid microtracer trial.
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