All information in this article is for educational purposes only. It is not for the diagnosis, treatment, prescription or cure of any disease or health condition.
AHVMA Journal • Volume 48 Fall 2017
TISSUE MINERAL ANALYSIS PATTERNS
IN 564 DOGS
By Ava Frick, DVM and Arlene Tolen, MPH, NC
ARL – Analytical Research Laboratory
CLIA – Clinical Laboratory Improvement Amendments
TMA - Tissue Mineral Analysis
Note: Standard elemental abbreviations are used throughout, for example Mg for Magnesium
Tissue Mineral Analysis (TMA) is a technique using soft
tissue hair or fur biopsy that provides a reading of the
mineral deposition in the cells and interstitial spaces of
the hair over a 2 to 3 month period. TMA can be used to
understand metabolism. It is another scientific measure that
can expand our understanding of health and processes that
impact illness in dogs. Mineral excess or deficiency is known
to produce certain physical and psychological symptoms.
The correlation of TMA results with clinical signs seen in
patients is discussed in this paper.
Tissue mineral levels and electrolyte patterns of calcium
(Ca), magnesium (Mg), sodium (Na), and potassium (K)
were analyzed in 564 dogs (300 male, 264 female; 99%
neutered or spayed) of variable breeds. Their ages ranged
from 1 to 15 years. Cases included all dogs presented to the
authors within a 12-year period.
Ninety-four percent (94%) of the 564 dogs were found to
have high Na to K ratios with low Ca and Mg. Ca, Mg, Na,
and K are prevalent minerals in the body, regulate osmotic
balance, and are involved in most body functions. According
to mineral analysis research, both results (levels and ratios)
are indicative of inflammation.
TMA is an accurate and scientifically valid approach to
assessing metabolic function and balance. TMA is responsive
not only to the trace mineral levels in the diet, but also to
all other factors which influence their metabolism including
stress, exercise, and endocrine and gastro-intestinal function.
Hair mineral analysis is a tissue mineral biopsy blueprint of
one’s biochemistry and the nutritional metabolic activity
that occurred during the period of hair formation. Hair
and fur are chemically indistinguishable, having the same
chemical composition, and are made of keratin. The primary
difference between hair and fur is the word usage. The hair
of non-human mammals is referred to as “fur,” while humans
are said to have hair. For the purposes of this paper, “hair”
refers to both hair and fur.
Hair is formed from clusters of matrix cells that make up
the follicles. During the growth phase, the hair is exposed to
the internal metabolic environment such as the circulating
blood, lymph, and extracellular fluids. As hair continues
to grow and reaches the surface of the skin, its outer layers
harden, locking in the metabolic products accumulated
during the period of formation. Hair analysis reveals
individual metabolic states in addition to the mineral status
of the individual animals (1, 2, 3).
Hair is 10-15% porous, and washing agents used in the
laboratory process can remove not only the exogenous
elements but also penetrate inside the shaft and wash
out some of the loosely bound minerals. Washing a
sample during test preparation removes quantities of
water-soluble macrominerals. When hair samples are not
washed by the laboratory prior to assessment, the analysis
can accurately measure macrominerals. Therefore, it
is important to select a laboratory that does not wash
the submitted specimen prior to the mass spectrometer
The laboratory will instruct the practitioner on the
protocol for collecting the sample from the animal (4-7).
For animals treated with topical insecticides and pesticides, it
is recommended to collect the sample during the last week of
the treatment cycle (i.e., 1 week before the next application
for most products). The animal can be bathed and clipped 4
hours later. Alternately, the ventrum, where the sample will
be collected, can be lightly sprayed with alcohol, wiped clean,
and then clipped, avoiding the topical application zone.
TMA is a non-invasive and a cost-efficient screening test (8).
It has been used throughout the world to assess individual or
herd nutritional status, providing a sensitive indicator of the
long-term metabolic trends from the effects of diet, stress, and
toxic metal exposure (9-14). Most deficiencies in animals are
brought about by altered relationships of minerals within the
body. It has become evident that both the retention and loss of
minerals by the animal are equally as important as the nutrients
consumed from the feed itself. This is valuable in determining
dietary needs as well as for recommending supplementation
Mineral ratios and levels can be changed by the
presence of toxic metals, nutritional deficiencies, infections,
illness, or stress from a myriad of sources (18, 19).
Assessment of an individual’s stage of stress and oxidation
rate (how fast the body metabolizes the food in exchange for
energy) can be determined by TMA (20). Measured stress
can come from external or internal sources. Helping or
treating stress with specific minerals can improve the health
status, resulting in better physical and emotional responses.
Many factors can influence the stress patterns of a TMA
Trends for common psychological conditions such
as depression, hyperactivity, anxiety, and mood swings may
be identified through TMA utilizing the levels and ratios of
calcium (Ca), magnesium (Mg), sodium (Na), and potassium
(K). Research confirms the intimate connection between
the body’s biochemistry and many emotional states (21-24).
Changes in body chemistry are reflected in the cellular
metabolism as measured by the hair tissue mineral
levels and inter-relationships found via TMA.
These can indicate metabolic dysfunction before clinical signs
occur. While blood values reflect what is in the blood,
hair analysis provides a record of how the body stores
and disposes of elements. The choice of hair as a testing
medium is based on the fact that blood chemistries
change dynamically from day to day, while hair values give
a more stable view of the overall mineral nutrition (25).
The purpose of this retrospective study was to show how
TMA has been used on a large dog population to establish
typical mineral patterns; to present why TMA is a valid test to
aid in nutritional assessment; to ascertain how these patterns
affect function, health and behavior; and to help determine
reasons why this can be occurring in dogs across the U.S.
MATERIALS AND METHODS
Tissue mineral levels and electrolyte patterns of Ca, Mg, Na
and K were analyzed in 564 dogs (300 male, 264 female; 99%
neutered or spayed) of variable breeds. Their ages ranged
from 1 to 15 years and included all dogs presented to the
authors within a 12- year period. The health conditions or
clinical signs of the dogs ranged from dermatitis, anxiety,
nervousness, aggression, gastrointestinal conditions,
metabolic conditions, neuromuscular, pain, and degenerative
joint disease. Some samples from apparently healthy dogs
were run for a baseline assessment by their owners (Table 1).
Table 1: Presenting Clinical Signs or Conditions
Presenting Clinical Signs or Conditions Number of Dogs
Included in Group
Anxiety, Aggression, Nervousness 302
Metabolic Conditions (diabetes, heart disease, liver
disease, kidney disease, cancer)
Degenerative Joint Disease 117
Apparently Normal 47
Seven groups of conditions or clinical signs that were seen in 517 of the total
group are listed here. Many of the dogs had multiple problems.
Another 47 dogs were apparently healthy without any complaints
from the owners. The TMA in those 47 was run as a baseline.
Their data is also included in the total statistics reported.
The fur was collected via clipping or cutting from the ventrum,
not longer than 1.25 cm in length and taken close to the skin.
Samples were weighed on a small scale provided by the laboratory
to at least 100 mg (less than 100 mg would produce insufficient
sampling), placed in a paper envelope, and submitted to the
Analytical Research Laboratory (ARL) in Phoenix, Arizona.
Computer-controlled mass spectrometers and induction-coupled
plasma (ICP) instruments are used today at hair testing
laboratories in the U.S. All commercial hair testing laboratories
in the U.S. are licensed and inspected annually by the federal
government as part of the Clinical Laboratory Improvement
Amendments (CLIA) act.
In this study, testing was performed
on a Perkin Elmer Elan 9000 ICP Mass Spectrometer. Results
are reported as mg % (parts per million times 10). The source
of standards for TMA used by the ARL is provided from original
dog research by Parmae Laboratory (Texas) along with 40 years
of research data collected by Dr. Paul Eck of ARL. A completed
report includes the optimal reference range, the patient’s level,
toxic metals, and mineral ratios (Table 2).
((sTable 2: Example of TMA report
There are thousands of biochemical reactions that ultimately
control metabolism, digestion, and the regeneration of
body tissues. The vast majority of these reactions depend on
minute amounts of trace minerals for their activity. If these
essential minerals are not present to fuel the processes, then
the body’s ability to regenerate, metabolize, or break down
noxious substances is compromised.
Because hair in the growth phase is exposed to the internal
metabolic environment (circulating blood, lymph, and
extracellular fluids) and retains the metabolic products
presented to it as it hardens, it becomes a perfect tissue sample
for testing body function, metabolic trends, and toxic metals.
TMA is a technique using soft tissue mineral biopsy that provides
a reading of the mineral deposition in the cells and interstitial
spaces of the hair over a 2 to 3 month period. This form of
testing allows for measurement capabilities of nutritional
alterations and improved recommendations for vitamin
and mineral supplementation or nutrient augmentation.
Ninety-four percent (94%) of the 564 dogs in the study
population had low Ca, Mg, and K, and high Na. This
correlates to a metabolic stress pattern, and the consistency of
these mineral patterns should not be ignored. Other studies
have reported similar findings; they linked this mineral
pattern to inflammation influencing T cells, cytokines, and
anti-inflammatory plasma proteins (26, 27). Another set
of parallel data confirmed that the deficiency of essential
mineral elements and Na overload can directly cause lipid
peroxidation and eventually hepatic damage (28).
Hair analysis offers a perspective for studying both
“disease” trends and for studying “stress” and physiological
coping mechanisms (29, 30). Disease trends and metabolic
changes can be documented and supported by using hair
Ca and Mg levels have been measured in patients with
fibromyalgia (34). Hair Ca concentration also correlates
to coronary heart disease (35). Levels of Ca and Mg have
been studied as part of the metabolic syndrome (36). Both a
deficiency and excess of these electrolytes have been shown
to be of pathogenic value in the development of endocrine
disorders and thyroid disease (37-40).
Nutrients can be unavailable for many reasons.
Malabsorption or bio-unavailability of specific minerals
can occur due to gut interference; improper form; lack
of other needed vitamins, minerals, or amino acids; and
ligand interruption, to name a few. Low taurine, low K,
and high Na all lead to low tissue Mg levels. Studies of both
animals and humans link minerals to healthy functioning
of the nervous system and behavior (41, 42). Nutrient
minerals must be available to support positive behavior.
Ca plays a fundamental role in metabolic processes, which
can be altered by small changes in extracellular ionized
Ca concentrations (43-46). A tissue Ca deficiency can
exhibit as a sympathetic dominant state (alarm or frightflight
reactions), anxiety, bruising, high blood pressure,
“fast” oxidation/metabolic state, insomnia, irritability,
muscle cramps and spasms, nervousness, hyperactivity,
osteoporosis, weakened ligaments and tendons, and tooth
decay (47,48). When the extracellular fluid level of Ca
ions drops below normal, the nervous system becomes
progressively more excitable because of increased
permeability of the neuronal membrane to Na (49).
A deficiency of tissue Mg affects the autonomic nervous
system, behavior, muscle tone and function (50-53). Clinical
signs include poor appetite, irritability, weakness, muscle
tremors, tetany, twitching, numbness, tingling, confusion,
disorientation, personality changes, apathy, memory loss,
skin lesions, tissue calcification, elevated cholesterol,
cardiovascular changes, tachycardia, elevated parathyroid
hormone, pancreatitis, and stress (42).
Magnesium deficiency increases catecholamine
secretion and sensitivity to stress, and may promote
aggressive behavior (54). Overall, Mg can act as a
neuron protector against aggression. Magnesium helps
to protect the nervous system locally and globally, is a
psycho-stabilizer, and plays an important role in aging
(55, 57). Increased catecholamine-induced intracellular
Mg loss is a causal factor in urinary loss of Mg (56).
When tissue K is deficient relative to Na, the body will tend
to develop hypertension, arrhythmias, and fatigue (57).
Excess Na will be able to enter the cell, which interferes with
cellular metabolism, especially protein synthesis. When this
happens, the cellular response is expressed as loss of energy
and manifests in the individual as exhaustion, submissive
behavior, and heightened anxiety or panic (58).
High tissue Na levels are indicative of excessive adrenal
gland activity and excitability. Excess Na combined with
too much phosphorus, low tissue levels of Ca, Mg, zinc
(Zn), and K result in adrenal insufficiency (59). Na,
when elevated in the tissue, is a stress that is associated
with anger and fear, the emotions of the “fight or flight”
response, and an indication of body inflammation (60).
With the activation of the stress response, the adrenal glands
increase their secretion of the hormone aldosterone, which
increases the retention of Na in the cells and tissues. As Na is
retained, Mg and Ca are lost from the cells and tissues. The
loss of Zn and Mg under stress appears to facilitate the stress
response by allowing for greater retention of Na (61, 62).
The primary mechanism by which psychological factors
predispose one towards a disease process is by means of the
stress response and its effect on nutrient minerals at the cellular
level. The stress response involves general systems responses
in which psychological, neurological, endocrine, and immune
system phenomena occur. Cellular energy production and
neuro-endocrine interference ensues. Mineral imbalance is a
factor in physiological stress and antisocial behavior (63, 64).
Stress has the capacity to impact minerals even more than
other nutrients. Zn and Mg are lost from the body’s tissue
reservoir in the “Acute Stage” of stress. Ca and copper are
lost in the “Resistance Stage” of stress. Loss of Na and K
are associated with the “Exhaustion Stage” of stress. Both
internal and external stressors create an increased need for
vitamins and minerals to recover and to prevent depletion
Dogs are carnivorous biased, which means they derive
optimal health and well-being by eating meat. Even though
they have omnivorous capacity and many manage to survive
on grain and carbohydrate-rich foods, a solely grainbased
diet is not the best choice for maximizing digestion,
absorption, health, and disease-free longevity.
Grains have been shown to interfere with Zn and
Ca assimilation (46). Eating carbohydrates lowers
Ca and Mg, and raises Na. Ca and Mg are sedative
elements, or known as the calming minerals. Lowering
their levels therefore increases the sensitivity of the
sympathetic division of the autonomic nervous system
and prepares the dog to respond to either fight or flight.
Mineral composition of the body is dependent not only
on food intake but on the efficiency or inefficiency of
neuroendocrine function (65). Hormones are known to
influence nutrients at several levels including absorption,
excretion, transportation, and storage. Nutrients in
turn can exert an influence on hormones (39, 66, 67).
The generations of dogs who were fed the dry grain-based
foods may have an impact on today’s canine
offspring that are now showing physiological imbalance.
Pesticides, herbicides, and insecticides are known to act as
endocrine disruptors in humans and animals. Herbicides
and fungicides affect Ca transport in plant mitochondria.
Herbicides and insecticides have been shown to interfere
with Ca metabolism in birds. Further, 354 drugs have been
reported to interact negatively with Ca, Mg, and Zn. Toxic
metals affect mineral metabolism and availability. Tannins
and polyphenols from tea, for example, reduce Mg in
tissue. These are all stressors that affect minerals (68, 69).
The majority (94%) of the 564 dogs studied were found to
have a high Na/K ratio with low Ca and Mg. According to
mineral analysis research, both results (the levels and the
ratio) are indicative of inflammatory processes and stress. This
correlates well with the clinical signs (general, physiological,
and psychological) reported in the cases studied.
Macrominerals are essential in metabolism in that they
activate catalytic and enzyme functions. Hair analysis is used
not only for measuring those minerals but also to monitor
the nutritional state of the dog until treatment benefits are
achieved and the effects of the treatment protocol have been
stabilized. The combination of feed ratio and hair analysis is an
invaluable screening tool to determine the correct program of
diet and supplementation for each dog's specific needs. TMA
is responsive not only to trace mineral levels in the diet but to
all other factors which influence their metabolism including
stress, exercise, and endocrine and gastro-intestinal function.
When performed to standards and correctly interpreted,
a TMA can be used as a screening tool in canine wellness
programs; and for those in suboptimal health, for monitoring
mineral deficiencies, mineral excesses, biochemical
characteristics, system imbalances, supporting behavior,
and endocrine indices (70-75). The use of hair TMA offers
a sophisticated and cost-effective approach (less than a
typical chemistry/CBC panel) that is very affordable for any
practitioner to incorporate into their nutritional assessment
There is currently nothing else available in the
way of a simple laboratory test that gives the veterinarian or
nutritional consultant a perspective of the internal nutritional
status of an animal. It can be a method to comply with AVMA
and AAHA recommendations regarding the importance
of offering annual or semi-annual nutritional evaluations.
This enables the veterinarian to present a nutritional
consultation that has significant value and direction toward
resolving health issues using individualized nutrition.
Technical assistance was recruited from Elaine Eisenbeisz of
Omega Statistics in Murrietta, California for the statistical
analysis. Manuscript submission preparation was enlisted
from Lisa Winstead in Rolla, Missouri.
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