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European Journal of Applied Sciences – Vol. 12, No. 4
Publication Date: August 25, 2024
DOI:10.14738/aivp.124.17502
Peregrina Lucano, A. A., Aguilar Uscanga, B. R., Solís Pacheco, J. R., & Rodríguez Arreola, A. (2024). Evaluating the Elemental
Composition of Mature Human Milk: Implications for Infant Health in the Guadalajara Metropolitan Area. European Journal of
Applied Sciences, Vol - 12(4). 522-535.
Services for Science and Education – United Kingdom
Evaluating the Elemental Composition of Mature Human Milk:
Implications for Infant Health in the Guadalajara Metropolitan
Area
Peregrina Lucano, Alejandro Aarón
Applied Pharmacokinetics Laboratory,
University Center of Exact Sciences and Engineering (CUCEI),
University of Guadalajara, Guadalajara, Jalisco, Mexico;
Aguilar Uscanga, Blanca Rosa
Human Milk Research Laboratory, Department of Farmacobiology,
University Center of Exact Sciences and Engineering (CUCEI,)
University of Guadalajara, Guadalajara, Jalisco, Mexico
Solís Pacheco, Josué Raymundo
Human Milk Research Laboratory, Department of Farmacobiology,
University Center of Exact Sciences and Engineering (CUCEI,)
University of Guadalajara, Guadalajara, Jalisco, Mexico
Rodríguez Arreola, Ariana
Human Milk Research Laboratory,
University Center of Exact Sciences and Engineering (CUCEI),
University of Guadalajara, Guadalajara, Jalisco, México
ABSTRACT
Environmental factors can significantly affect the health of infants through human
milk, including the presence of toxic metals from various sources. In the
metropolitan area of Guadalajara, current contamination levels are particularly
concerning. Consequently, a study was conducted using Inductively Coupled
Plasma Mass Spectrometry to analyze mature human milk from 30 housewives
with various health issues. The results showed concentrations of toxic elements
such as Pb, Zn, Cu, Hg, Cr, and as below 0.01 μg/L, while Al was above 0.51 μg/L
and Cd above 0.7 μg/L. Additionally, variations were observed in essential
elements such as Na, Mg, K, and Ca, with a notably high Na ratio in all cases, same
case in Rb. In conclusion, this study revealed the presence of low concentrations of
toxic metals in the breast milk of women from the metropolitan area of
Guadalajara, except for Al and Cd. Significant variations in essential elements were
also found, particularly a high Na ratio, which underlines the need to monitor
environmental contaminants that may affect infant health through breastfeeding
and identify their origin.
Keywords: human milk, toxic metals, contamination, infant health.
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Peregrina Lucano, A. A., Aguilar Uscanga, B. R., Solís Pacheco, J. R., & Rodríguez Arreola, A. (2024). Evaluating the Elemental Composition of
Mature Human Milk: Implications for Infant Health in the Guadalajara Metropolitan Area. European Journal of Applied Sciences, Vol - 12(4). 522-
535.
URL: http://dx.doi.org/10.14738/aivp.124.17502
INTRODUCTION
According to the World Health Organization (WHO), breastfeeding is the ideal food for the
child exclusively during the first six months of life and beyond these months alongside other
foods, as it is essential for children to reach their full potential in terms of growth, health, and
development. However, the fat and protein content of breast milk can make it a route of
excretion and a vehicle for the passage of toxic substances to the child, such as inorganic
elements and perfluorinated compounds, including toxic metals [1]. Studies conducted on
pediatric populations have demonstrated that metals such as cadmium, lead, mercury, copper,
zinc, and arsenic are potentially toxic, with hemotoxic, neurotoxic, and nephrotoxic effects
even at low blood concentrations. Lead, a highly harmful metal, can accumulate in the body
over time and affect multiple systems. It has been linked to cognitive deficits, behavioral
problems, and developmental delays in children exposed during critical periods of growth and
development [2, 3]. In the case of arsenic, which is naturally present in water and soil, it can
contaminate food and water sources. Prolonged exposure to this element has been associated
with a higher incidence of diverse types of cancer, cardiovascular diseases, and adverse
reproductive outcomes. On the other hand, Cadmium, primarily released into the
environment through industrial processes, can accumulate in the body and have adverse
effects on the kidneys, respiratory system, and skeletal system. Long-term exposure to
cadmium is linked to conditions such as osteoporosis, kidney damage, and lung diseases.
These are just some of the adverse effects that may occur [4, 5].
Exposure to toxic metals has a significant impact on biomolecules, particularly proteins with
enzymatic activity. These proteins play a crucial role in numerous biological processes,
including the regulation of metabolism, immune response, and cell repair. Such exposure can
alter the structure and function of certain proteins, leading to impaired enzymatic activities
and, consequently, the normal functioning of biological systems. This disruption can trigger
multisystemic pathology, which is a critical aspect for understanding the bodily distribution of
these metals, as well as the mechanisms of their elimination and excretion [6, 7]. The main
systems affected by exposure to toxic metals include the gastrointestinal, central and
peripheral nervous, and hemic systems. In the gastrointestinal tract, these metals interfere
with nutrient absorption. In the neurological system, they can cross the blood-brain barrier,
affecting brain function and causing cognitive impairment (such as in cases of lead and
mercury exposure), neurological developmental disorders, and neurodegenerative diseases.
The hematopoietic system is compromised in terms of blood cell production and the onset of
anemia. Additionally, elements like arsenic and cadmium can increase the risk of developing
cancer in various tissues and organs. To adequately understand and address the effects of
toxic metals on the body, it is essential to consider their multisystemic pathology and the
most affected systems. This understanding will enable the development of strategies to
prevent exposure, detect and treat associated diseases, and implement effective measures to
eliminate and excrete these metals from the body [6, 8, 9].
The population can be exposed to these contaminants due to their widespread dissemination
in the environment [10]. Exposure to toxic metallic elements can occur in the environment air,
water, and soil particularly during occupational activities, as well as through the consumption
of contaminated food [11]. In the case of breastfeeding and/or pregnant women, exposure has
a more severe impact. Once heavy metals and metalloids enter the human body, their affinity
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for proteins allows them to mobilize within the organism, travelling through red blood cells or
other plasma components until they accumulate in various organs and tissues, including
bones, teeth, nails, and hair. These toxic agents can bioaccumulate in the maternal body and
subsequently transfer to the infant during pregnancy and breastfeeding, directly affecting the
health of the mother-child pair [12]. Through the method Inductively Coupled Plasma Mass
Spectrometry (ICP-MS), this study aimed to evaluate the levels of 29 elements: Li, Be, Na, Mg,
Al, K, Ca, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, Ga, As, Se, Rb, Sr, Ag, Cd, In, Cs, Ba, Hg, Pb, Bi, U, in mature
human milk from women residing in the metropolitan area of Guadalajara. The objective was
to assess exposure to these elements and thereby identify potential health implications.
MATERIAL AND METHODS
Human Milk Collection
The concentrations of the 29 elements in mature breast milk were determined from 30
human milk samples collected from healthy primiparous and multiparous lactating mothers,
as well as from diabetics, smokers, drug addicts, with preeclampsia and syphilis in the
metropolitan area. from Guadalajara. These women are nursing assistants at the Old Civil
Hospital of Guadalajara “Fray Antonio Alcalde” in Jalisco, Mexico. According to data provided
by the hospital, the age of the mothers ranged between 14 and 32 years, all of them reported
being housewives. Three of them mentioned being smokers and two admitted to having
consumed some type of drug while breastfeeding. None of the mothers analyzed were
occupationally exposed to metals. The project received approval from an Ethics Committee
(HCG/CEI-0907/22 and research registration 141/22 approved on June 8, 2022), and
informed written consent was obtained from all human milk donor mothers in accordance
with the ethical principles outlined in the Declaration of Helsinki of 1964, revised in 2008, and
the regulations of the General Health Law on Health Research 2004, last reformed in 2008.
Human milk collection was performed using a manual pump after sanitizing the donor
mothers. The milk was then deposited in special polypropylene bags designed for BPA-free
breast milk with a wide neck, pre-sterilized for immediate use, with a capacity of up to 6 oz,
and equipped with a double anti-drip safety closure system for safe storage. These bags
feature reinforced edges and a double layer for secure refrigeration or freezing, while also
undergoing verification of organoleptic properties (smell, color) and possible contamination
to ensure authenticity. After collection, the samples were stored in a cooler and transported to
the Pharmacokinetics laboratory for further analysis.
Analysis of Metals in Human Milk:
For determination of elements in human milk, it was used an equipment CEM brand
microwave oven model MARS-5, Agilent Technologies model 7800 Inductively Coupled
Plasma Mass Spectrometer (ICP-MS) with Agilent Technologies SPS 4 autosampler and a 5%
ICP-MS HNO3 multi-element calibrator standard Perkin Elmer Pure Plus® with a
concentration of 10 μg/mL was used of Li, Be, Na, Mg, Al, K, Ca, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, Ga,
As, Se, Rb, Sr, Ag, Cd, In, Cs, Ba, Hg, Pb, Bi and U.
Preparation of Human Milk Samples:
Sample preparation involved digestion with nitric acid (60-70%) Plasma PURE®, which was
heated in a microwave oven. A volume of 0.5 mL of the liquid human milk sample was
measured using a micropipette and placed in PFA Teflon tubes with a 25-mL capacity screw
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Ag y=5.837x+1.081 0.999
Al y=9.095x+6.007 0.999
As y=4.320x+8.008 0.999
Ba y=1.461x+1.972 0.999
Be y=4.180x 0.999
Bi y=2.053x+5.477 1
Ca y=4.112x+3.304 0.998
Cd y=1.201x+1.502 0.999
Co y=4.384x+3.203 0.999
Cr y=2.735x+4.004 0.999
Cs y=8.439x+6.107 0.999
Cu y=2.974x+2.950 0.999
Fe y=2.470x+2.408 0.998
Ga y=1.218x+6.006 0.999
Hg y=3.196x+1.001 0.999
In y=9.475x+1.392 0.999
K y=2.288x+3.597 0.998
Li y=3.968 x + 2.002 0.999
Mg y=2.072x+1.047 0.995
Mn y=1.845x+9.210 0.999
Na y=3.828x + 3.392 0.999
Ni y=1.155x+3.284 0.998
Pb y=1.435x+7.089 0.999
Rb y=2.223x+3.103 0.999
Se y=3.828x+2.002 0.999
Sr y=3.099x+7.330 0.997
U y=2.462x+9.313 1
V y=2.238x+4.012 0.998
Zn y=5.972x+2.051 0.998
Milk samples in our experiment exhibited highly variable concentrations of elements
including Na, Mg, Al, K, Ca, Fe, Cu, Zn, Rb and Cd, as shown in Table 4. The results obtained for
sodium did not show significant differences in the concentrations of this element (p>0.05)
among women with different health profile. The values for K were significantly different
across all health profiles (p<0.05), with women with syphilis showing the highest
concentrations compared to the other health profiles evaluated. In the case of Mg, there was a
significant difference (p<0.05) between healthy women and those with diabetes, drug
addiction, elevated triglycerides, and smokers. Additionally, women with syphilis showed
significant differences compared to women with drug addiction and smokers. No significant
differences (p>0.05) were found in the concentrations of Al or Fe among the study population,
except between healthy women and those with syphilis (for both Al and Fe), and between
women with syphilis and those with diabetes (for Fe) (p<0.05). A significant difference
(p<0.05) in Cu concentrations was found between diabetic women and those with syphilis,
between healthy women and drug users, as well as between healthy women and smokers.
There is a significant difference (p<0.05) in Zinc concentrations between healthy women and
those who are drug users, syphilitic, and smokers. In the case of Cd, it was found only in
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Peregrina Lucano, A. A., Aguilar Uscanga, B. R., Solís Pacheco, J. R., & Rodríguez Arreola, A. (2024). Evaluating the Elemental Composition of
Mature Human Milk: Implications for Infant Health in the Guadalajara Metropolitan Area. European Journal of Applied Sciences, Vol - 12(4). 522-
535.
URL: http://dx.doi.org/10.14738/aivp.124.17502
smokers, one diabetic woman, one woman with syphilis, and drug addicts. There is a
significant difference (p<0.05) between smoker women regarding diabetics, drug addicts and
those with syphilitic. The exposition to Cadmium has been recognized when it was discovered
in Itai-itai disease was caused by the consumption of cadmium-contaminated rice in 1955.
Since that finding, numerous studies have documented the effects exposure of this element on
the health of the general population, even in the absence of specific industrial exposure, with
food and tobacco smoke being the primary sources of exposure. This could explain why the
smoking women in this study had cadmium in their milk. Additionally, environmental factors
such as contamination may contribute to the variability observed between samples obtained
from the same population and when compared with samples analyzed in studies conducted in
other countries. Furthermore, the influence of environmental agents such as contamination,
exposure to toxic substances, and geographic variations may contribute to the observed
differences Cd element concentrations in breast milk samples from different populations and
regions [13, 14].
Table 4: Elements found in human milk samples in μg/L (Mean ± Standard Deviation)
23 Na 24 Mg 27Al 39K
44Ca 56Fe 63Cu 66Zn 85Rb 114Cd
Sample ID Observations (M±SD)
L1 healthy 171.68±2.35 24.62±0.09 0.64±0.02 481.22±1.06 105.00±0.54 1.28±0.18 4.34±0.04 0.96±0.00
L2 healthy 404.25±3.78 30.72±0.37 360.84±4.68 79.97±1.44 0.64±0.04 0.56±0.03
L3 healthy 123.20±1.06 36.52±0.79 347.11±0.72 83.81±1.08
L4 healthy 153.15±3.86 31.06±0.08 482.68±0.20 81.64±0.63 3.70±0.02 0.64±0.01
L5 healthy 119.81±4.07 19.53±0.54 535.94±16.79 74.18±1.78 1.96±0.14 0.79±0.02
L6 healthy 211.14±2.28 28.06±0.76 0.45±0.12 654.52±1.32 99.10±0.36 3.28±0.06 1.20±0.01
L7 healthy 378.26±0.58 25.31±0.08 0.71±0.02 364.59±1.55 45.24±1.08 1.19±0.0 1.14±0.04 0.55±0.00
L8 healthy 136.32±1.09 18.80±0.12 1.00±0.01 436.36±1.53 57.08±3.00 1.15±0.00 2.98±0.03 0.72±0.00
L9 healthy 779.08±2.67 26.42±0.03 0.56±0.07 349.04±7.38 78.67±2.50 1.57±0.08
L10 healthy 729.27±27.54 30.17±1.19 1.15±0.01 484.72±21.35 52.72±2.13 1.72±0.07 1.19±0.18 0.70±0.02
L11 healthy 122.85±0.85 29.25±0.10 0.05±0.05 302.95±0.95 71.32±0.32
L12 healthy 299.50±1.75 25.49±0.23 2.25±0.10 505.39±4.97 76.51±0.88 0.81±0.01
L13 healthy 97.21±1.65 32.55±0.03 457.54±0.49 72.04±1.40 0.70±0.02 0.76±0.03 0.95±0.01
L14 healthy 93.62±0.62 28.16±0.43 363.00±4.6 61.99±1.05 0.80±0.01
L15 healthy 68.09±0.20 28.28±0.25 0.69±0.06 379.54±1.53 70.45±0.91 0.76±0.04 0.73±0.01
L16 healthy 83.89±0.20 24.57±0.43 339.50±0.57 52.10±1.10 1.36±0.02 0.69±0.05 0.54±0.01
L17 healthy 45.60±0.05 26.39±0.38 345.97±2.11 55.51±1.78 0.55±0.02
L18 healthy 99.31±1.89 21.56±0.40 1.07±0.05 397.61±6.83 85.28±4.88 1.67±0.02 0.63±0.02
L19 healthy 532.47±1.74 31.44±0.14 431.77±1.28 64.22±0.58 0.75±0.01
L20 healthy 505.63±5.95 33.02±0.17 0.58±0.05 490.01±3.65 89.11±1.20 1.59±0.03 2.55±0.12 0.64±0.01
L21 healthy 123.35±2.86 28.05±0.72 472.72±7.38 53.01±1.77 1.59±0.03 2.29±0.11 0.84±0.02
L22 healthy 404.19±2.48 31.83±0.37 0.50±0.07 439.85±3.33 72.99±0.69 0.89±0.01 3.48±0.08 0.79±0.01
L23 healthy 610.61±7.27 31.18±0.22 434.59±6.63 58.55±1.77 0.79±0.02 0.73±0.01
L24 healthy 195.89±1.96 25.54±0.12 590.17±0.21 62.11±2.11 0.69±0.02 3.08±0.05 0.68±0.01
L25 healthy 321.40±0.81 24.48±0.14 480.37±4.97 56.89±0.01 0.68±0.01 1.56±0.04 0.61±0.02
L26 healthy 410.73±3.33 24.35±0.10 384.94±0.43 65.99±1.23 0.71±0.01 1.24±006 0.59±0.01
L27 healthy 309.52±1.12 40.20±0.08 0.56±0.03 457.59±2.72 75.31±1.66 1.41±0.02 0.69±0.01 0.62±0.02
L28 healthy 79.82±2.29 21.09±0.63 327.85±18.31 72.55±1.96 1.21±0.13 0.67±0.04
L29 healthy 107.50±2.93 22.9±0.72 325.06±10.50 59.52±0.06 0.55±0.01
L30 syphilis 83.87±1.45 18.88±0.30 324.04±1.80 68.34±0.34 0.51±0.01
L31 syphilis 1266.23±30.74 10.05±0.18 162.97±0.65 61.50±0.39 1.01±0.05
L32 syphilis 1389.63±3.37 28.86±0.20 1496.63±26.36 15.37±0.13 0.83±0.26 3.76±0.05 1.83±0.01
L33 syphilis 490.66±4.48 61.10±0.41 0.52±0.02 1064.62±11.25 80.74±0.17 0.56±0.01 1.22±0.01 5.95±0.07 1.51±0.01 1.7±0.02
L34 syphilis 272.24±3.48 40.57±0.20 838.57±7.52 52.14±1.47 0.91±0.01 7.76±0.03 1.20±0.02
L35 diabetic 728.06±5.69 57.54±0.89 0.51±0.06 1242.14±14.17 82.28±0.60 0.51±0.04 2.61±0.01 7.96±0.08 2.16±0.04
L36 smoker 838.26±5.63 62.86±0.16 1141.09±19.44 68.31±3.76 0.60±0.01 1.25±0.01 5.45±0.02 1.59±.0.01 0.9±0.02
L37 smoker 230.10±1.32 36.74±0.03 953.13±20.24 38.11±0.08 0.66±0.01 5.94±0.02 1.65±0.01 0.8±0.04
L38 smoker 392.68±4.48 60.59±0.43 1055.98±9.18 57.82±1.58 1.78±0.04 0.88±0.02 4.04±0.03 1.64±0.03 0.7±0.11
L39 High tgl 158.91±0.52 50.43±0.44 733.70±3.52 47.07±0.93 0.94±0.01 3.89±0.02 1.29±0.01
L40 drug addict 623.76±2.49 45.08±0.40 902.48±3.11 41.71±0.54 0.95±0.03 0.74±0.01 3.68±0.05 1.36±0.01
L41 drug addict 565.43±0.89 59.10±0.19 1147.82±14.68 72.14±4.32 1.21±0.01 5.54±0.02 1.79±0.01 0.7±0.09
L42 diabetic 244.72±1.25 33.15±0.63 0.87±0.01 522.05±10.79 32.77±0.95 0.58±0.05 0.69±0.01 2.44±0.03 0.88±0.01 0.9±0.1
L43 diabetic 188.06±0.31 23.89±0.39 449.22±3.71 39.45±0.75 1.72±0.02 0.79±0.01
L44 diabetic 269.21±0.78 37.75±0.25 474.86±0.46 38.71±0.93 0.71±0.02 1.17±0.01 0.81±0.1
L45 preeclampsic 187.56±0.03 30.01±0.49 595.33±1.82 27.89±0.77 2.21±0.01 0.99±0.01
Concentrations of metals such as As, Pb, that have been widely considered potentially toxic to
humans [15, 16,] were determined at concentrations <0.01 μg/L. This result is consistent with
previous research indicating low concentrations of arsenic in human milk [17]. The presence
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Peregrina Lucano, A. A., Aguilar Uscanga, B. R., Solís Pacheco, J. R., & Rodríguez Arreola, A. (2024). Evaluating the Elemental Composition of
Mature Human Milk: Implications for Infant Health in the Guadalajara Metropolitan Area. European Journal of Applied Sciences, Vol - 12(4). 522-
535.
URL: http://dx.doi.org/10.14738/aivp.124.17502
in breast milk samples from smoking mothers, drug-addicted mothers, and those with
elevated triglycerides, compared to healthy mothers. Research indicates that the
concentrations of sodium (Na) and potassium (K) in human milk undergo significant changes
during lactation, particularly in the first week postpartum. Specifically, sodium concentrations
tend to decrease rapidly. However, the effects of tobacco and recreational drug use, as well as
health issues such as diabetes and high triglycerides, on these elements have not been widely
studied [13].
The influence of tobacco and recreational drug use on Na, K, Ca, and Mg concentrations in
breast milk is likely multifactorial. Tobacco smoke and certain drugs can alter electrolyte
balance and affect ion transport mechanisms in the mammary gland. Additionally, these
substances can impact general metabolism and nutrient absorption, indirectly affecting breast
milk composition. Health conditions such as diabetes and high triglycerides involve hormonal
and metabolic changes that can potentially influence electrolyte concentrations in breast milk.
Alterations in hormone levels and metabolic processes can disrupt the homeostatic regulation
of Na, K, Ca, and Mg [25, 26].
The Na ratio is considered a marker for the process of secretory activation of milk. An
elevated concentration in human milk may indicate increased permeability of the mammary
epithelium, related to inflammatory processes such as mammary engorgement or mastitis.
The balance of sodium and potassium in breast tissue is essential to maintaining the integrity
and proper function of the breast epithelium. Potassium is the main intracellular cation, while
sodium is found in higher extracellular concentrations. Normally, there is an electrochemical
gradient that keeps potassium inside cells and sodium outside cells. However, during
inflammatory processes in the mammary gland, such as engorgement (excessive
accumulation of milk) or mastitis (breast infection), an increase in mammary epithelium
permeability can occur. This allows increased sodium entry into mammary cells and a
corresponding decrease in intracellular potassium [27]. The resulting imbalance in the Na
ratio can affect mammary cell function and compromise homeostasis. Increased sodium
uptake can lead to disturbances in osmotic balance and ionic gradients, affecting mammary
cell function and health. An elevated Na ratio in human milk may signal inflammation in the
mammary gland. Breast engorgement and mastitis involve a localized inflammatory response,
with mediators and cytokines released that affect tissue permeability and milk composition.
Importantly, an elevated Na ratio in human milk should not be considered a definitive
diagnosis of engorgement or mastitis. Clinical evaluation and other tests are necessary to
confirm these conditions. However, observing an unbalanced Na ratio in milk may warrant
investigating potential inflammatory processes in the mammary gland [28].
Our study found aluminum (Al) concentrations of up to 2.25 μg/L, contrasting with some
studies [29, 30], where Al was not quantifiable, while in another study [31], our findings were
lower. To date, it remains unclear if Al has any biological function in the human body.
However, it is well known as a contaminant in food and pharmaceuticals. Its presence in the
body has been linked to the development of Alzheimer's disease, raising concerns that its
presence in breast milk could have harmful effects on infant health. Humans are exposed Al
through many factors as diet, vaccine adjuvants, antacids, and, with Al-based antiperspirants
significantly increasing local exposure in the under area [32]. This region, particularly the