BMB Rep.

2015; 48(4): 209-216

BMB www.bmbreports.org
Reports
Invited Mini Review

Technical and clinical aspects of cortisol as a biochemical

marker of chronic stress
Do Yup Lee , Eosu Kim * & Man Ho Choi *
1 2, 3,

1
Department of Bio and Fermentation Convergence Technology, Kookmin University, Seoul 136-702, 2Department of Psychiatry, Institute
of Behavioral Science in Medicine, Brain Korea 21 Plus Project for Medical Science, Yonsei University College of Medicine, Seoul 120-752,
3
Future Convergence Research Division, Korea Institute of Science and Technology, Seoul 136-791, Korea

Stress is now recognized as a universal premorbid factor through situations like exams or work deadlines. However, an
associated with many risk factors of various chronic diseases. extreme amount of stress can lead to negative consequences
Acute stress may induce an individual’s adaptive response to and adversely affect the immune, cardiovascular, neuro-
environmental demands. However, chronic, excessive stress endocrine, and central nervous systems (1). In particular, chron-
causes cumulative negative impacts on health outcomes ic stress can have a serious impact due to sustained high levels
through “allostatic load”. Thus, monitoring the quantified of the chemicals released in the “fight or flight” response, which
levels of long-term stress mediators would provide a timely involves endocrine system releasing glucocorticoids (2, 3).
opportunity for prevention or earlier intervention of stress- 　Cortisol, which is synthesized from cholesterol, is the main
related chronic illnesses. Although either acute or chronic glucocorticoid in the zona fasciculate of human adrenal
stress could be quantified through measurement of changes in cortex. Its secretion in response to biochemical stress contrib-
physiological parameters such as heart rate, blood pressure, utes to the well-characterized suppression of the hypo-
and levels of various metabolic hormones, it is still elusive to thalamic-pituitary-adrenal (HPA) axis on health and cognition
interpret whether the changes in circulating levels of stress events (4-6). Since the vast majority of cortisol actions rely on
mediators such as cortisol can reflect the acute, chronic, or binding to cytosolic receptors, only a small fraction of un-
diurnal variations. Both serum and salivary cortisol levels bound, free cortisol is revealed to be biologically active. It
reveal acute changes at a single point in time, but the overall comes out of the mitochondrion, migrates out of the cell into
long-term systemic cortisol exposure is difficult to evaluate due the extracellular space and into the bloodstream. Due to its
to circadian variations and its protein-binding capacity. Scalp low molecular weight and lipophilic nature, unbound cortisol
hair has a fairy predictable growth rate of approximately 1 enters the cells through passive diffusion, which makes it fea-
cm/month, and the most 1 cm segment approximates the last sible to measure the free cortisol in many body fluids (7).
month’s cortisol production as the mean value. The analysis of 　In general, cortisol levels in blood increase during the early
cortisol in hair is a highly promising technique for the retro- morning (highest at about 8 a.m.) and decrease slightly in the
spective assessment of chronic stress. [BMB Reports 2015; evening and during the early phase of sleep (8). The timing of
48(4): 209-216] blood sampling is therefore very important. While its assess-
ment in sweat or tears is only of theoretical importance and
urinary cortisol of decreasing interest, salivary cortisol may
INTRODUCTION have some advantages over the assessment of cortisol in blood
(9, 10). Since the hormone levels in biological fluids fluctuate
Stress can lead to both physical and psychological health on a daily basis, cortisol extracted from the hair fiber has been
issues. Some stress can be beneficial at times by producing a investigated (11-13). This review discusses on the methods in-
boost that provides the drive and energy to help people get volved in mass spectrometry-based metabolomic studies for
identification of biomarkers in chronic stress, which is more
focused on hair cortisol. Comparative statistical analyses of
*Corresponding authors. Eosu Kim, Tel: +82-2-2228-1620; Fax:
crucial aspects are also included to facilitate the understanding
+82-2-318-0891; E-mail: kimeosu@yuhs.ac, Man Ho Choi, Tel:
+82-2-958-5081, Fax: +82-2-958-5059, E-mail: mh_choi@kist.re.kr of recent advances in the metabolic platform on mining
biomarkers.
http://dx.doi.org/10.5483/BMBRep.2015.48.4.275
STRESS AND THE ADRENAL GLAND
Received 17 October 2014
The two adrenal glands are located on top of the kidneys, and
Keywords: Allostasis, Cortisol, Hair, Mass spectrometry, Metabolo-
mics, Stress
these glands produce hormones in response to stress. Each

ISSN: 1976-670X (electronic edition)

Copyright ⓒ 2015 by the The Korean Society for Biochemistry and Molecular Biology
This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/li-
censes/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

adrenal gland consists of a central area, called the medulla, an increase of blood pressure and blood volume. Glucocorti-
and an outer area of the cortex (Fig. 1). In case of the apparent coids promote fat and protein breakdown and glucose syn-
threat, the hypothalamus sends direct signals via the sym- thesis. Cortisol is the major glucocorticoids, and it regulates or
pathetic nervous system to the adrenal glands, causing them to supports a variety of important cardiovascular, metabolic, im-
release a catecholamine and epinephrine (same as adrenaline). munologic, and homeostatic functions (1).
It leads to an urgent action by stimulating faster breathing and
heart rates. The adrenal medullar also secrete another cat- ALLOSTASIS AND ALLOSTATIC LOAD
echolamine, norephinephrine, which works with epinephrine
to stimulate liver cells to release glucose to make more fuel The term, stress, was originally adopted from engineering
available for cellular respiration. These hormones have (measure of the internal forces induced by deformation of a
short-term effects as the nerve impulses are sent from the body), but it is now referred to ‘threats or anticipation of
hypothalamus. Due to the short half-life of blood catechol- threats to an organism’s homeostasis’ (15). Thus stress events
amine, meticulous care must be taken to obtain blood samples could be understood as any stimuli that cause alterations in
consistently vis-à-vis the stress immersion experience (14). homeostasis for adaptation to the environment. These changes
　Finding a “gold standard” biomarker for chronic stress has in homeostasis are referred to as ‘allostasis’, which can be ex-
been proven to be challenging, given its complex etiology and emplified by increased heart rate or blood pressure and en-
highly individual manifestations, while the biomarkers of acute hanced systemic metabolism. In general, allostasis can be
stress have been well-defined and are primarily used to assess adaptive or maladaptive depending on its degree or contextual
the release of catecholamine. Hormones secreted by the adre- relevance; mediators of allostasis, such as metabolic hor-
nal cortex provide a slower, longer-acting (chronic) response mones, could contribute to healthy adaptation and pathophysi-
to the stress. In this event, the hypothalamus secretes a releas- ology (16). The concept of ‘allostatic load’ indicates an altered,
ing hormone which causes the anterior pituitary to secrete an ‘new set point’ of homeostasis, resulting from cumulative ef-
adrenal-stimulating hormone, adrenocorticotrophic hormone fects of allostatic responses which are chronic, excessive, or
(ACTH); and this signals the cells in the adrenal cortex to pro- poorly regulated (15, 16). For example, the increased serum
duce and secrete corticosteroids. Among them, mineralocorti- glucose level is responsible for a single acute stressful event,
coids, like aldosterone, can regulate water and sodium re-ab- which can be called as ‘allostatic response’. Also, diabetes
sorption in the kidneys. It also regulates the active secretion of (insulin resistance) resulting from repetitive chronic stress can
potassium in the principal cells of the cortical collecting tubule be understood as an ‘allostatic overload’, in which the base-
and protons via proton ATPases in the luminal membrane of line fasting glucose level has been newly set to a higher level
the intercalated cells of the collecting tubule, which results in than before (Fig. 2). Therefore, the biomarkers of allostatic

Fig. 1. The differential stress hor-

mones secreted by adrenal cortex and
medullar. Catecholamines cause gen-
eral physiological changes that pre-
pare the body for physical activity
(fight-or-flight response) in the short-
term response. Some typical effects in-
clude increases in heart rate, blood
pressure, and blood glucose levels,
and other general reactions of the
sympathetic nervous system. Corticoids
are involved in a wide range of phys-
iological processes including chronic
stress response, immune response, and
regulations of inflammation, carbohy-
drate metabolism, protein catabolism,
blood electrolyte levels, and behavior.
Data are taken from “https://www.
studyblue.com/notes/note/n/07-adrenal-
glands/deck/1109539”.

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 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

potential of utilizing cortisol as a biomarker for various chronic

illnesses or their pre-disease states (21, 22). Currently, it seems
that such role of cortisol, as one of biomarkers for the allostatic
load, may be largely accepted (6). However, there are still con-
troversial issues regarding the sampling and measurement
methods of these hormones that are fluctuated by environ-
mental contexts as well as by the circadian cycle. However,
the most important issues might be the possibility that the level
measured once at a current point could reflect the past history
of an individual’s stress load, just as HbA1c indicating the de-
gree of ‘glucose load’ for the past 3 months. The level of corti-
sol in scalp hair is now considered as a promising biomarker
Fig. 2. Stress, allostasis, and allostatic load. Stress is any stimulus
inducing either adaptive or maladaptive allostasis (changes in ho-
for assessing the averaged level of one’s past stress burden dur-
meostasis) of stress mediators, which constitute the autonomic ing a given period, and it will be further discussed in this
nervous system (blood pressure, catecholamines), metabolic hor- paper. Its utility has been demonstrated across diverse clinical
mones (cortisol, insulin), and pro- & anti-inflammatory cytokines. settings from neonates to old age, indicated by associations of
If stress stimuli are excessive and repetitive, recovery to the origi-
nal homeostatic levels may be incomplete (indicated by the sec-
hair cortisol levels with babies’ stress in the neonatal intensive
ond blue arrow). As a result, chronic stress can make a body sys- care unit (23), children’s stress at school entry (24), and vari-
tem to anticipate, as if such a new (stressful) environment would ous metabolic and neuropsychiatric disorders including acute
persist, demanding a newly defined set point for future adap- myocardial infarction (25), heart failure (26), metabolic syn-
tation. Thus, the difference between the new and old set points
can be understood as a ‘cumulative burden of adaptation to
drome (27), and post-traumatic stress disorder (28).
stress’- i.e., allostatic load. Examples of allostatic load may be
found in the primary mediators (hypercortisolemia, increased infla- ADVANCES IN CORTISOL DETECTION IN
mmatory cytokines), secondary outcomes (elevated blood pressure, METABOLITE PROFILING
overweight, insulin resistance), or tertiary outcomes (hypertension,
diabetes, obesity, coronary heart disease, neurodegenerative dis-
orders). For more details, see reference # 6. Because more than 90% of circulating cortisol in human se-
rum is protein-bound, changes in the binding proteins can al-
ter the levels of serum total cortisol without influencing the
load, if available, could be used for measuring and predicting free concentrations of cortisol. Total cortisol could be mislead-
the cumulative biological risks of impending illnesses, and ingly lower than anticipated, resulting in the incorrect inter-
they are ideal indicators for detecting the ‘pre-phase’ of ill- pretation that adrenal function is impaired. This is important
nesses (6, 17, 18). because the current standards for defining normal adrenal
　Mediators of allostasis constitute the autonomic nervous sys- functions are based on healthy people who have normal levels
tem (catecholamines), metabolic hormones, and various cyto- of binding proteins. Measuring serum free cortisol levels in
kines (15, 16). Among these, cortisol is the one that has been critical illnesses may help to prevent the unnecessary use of
paid great attention based on the concept of ‘glucocorticoid glucocorticoid therapy (21). Although the free cortisol hypoth-
cascade hypothesis’ (19). This hypothesis explains how and esis has been widely used, it has also been suggested that cor-
why the cortisol actions could be related to pathophysiology tisol binding globulin (CBG)-bound cortisol may have physio-
upon overload of stress. Psychological and physical stresses in- logical effects on target tissues (22).
crease the circulating cortisol levels. In the acute state, in- 　There is a high correlation between salivary cortisol levels
creased cortisol induces adaptive responses via enhancing ca- and unbound cortisol in plasma and serum, which remains
tabolic processes to supply more energy to the body. In gen- high during the circadian cycle and under different dynamic
eral, increased cortisol levels return to the basal levels by feed- test such as ACTH stimulation (22, 29). Since free cortisol rep-
back inhibition mechanisms through the hypothalamus, pre- resents the biologically active hormone fraction, salivary corti-
frontal cortex, and most importantly, hippocampus (20). When sol measures have early been considered as a better method
stressful stimuli are repeated chronically, circulating cortisol is than serum cortisol for the evaluation of adrenocortical func-
maintained at higher levels over a prolonged period. Chronically tion (9). Recently, the late-night salivary cortisol has been
elevated cortisol levels now causes damages on hippocampal showing a superior diagnostic performance as the primary bio-
and cortical neurons (19), which are the main regions where chemical diagnostic test for Cushing’s syndrome (10). Cortisol
the feedback inhibition starts. As a result, even when stress in biological fluids has been extensively evaluated with corti-
stimuli disappear, cortisol levels could be maintained at higher sone, which may reveal the activity of 11-hydroxysteroid de-
levels beyond the physiologically normal range due to a vicious hydrogenase (11-HSD) by mass spectrometry-based metabo-
cycle caused by the already damaged feedback mechanism. lite profiling techniques (30-32). These analytical methods
　Consequently, a great number of studies have explored the have shown the acceptable analytical sensitivity and the se-

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 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

lectivity in trace amounts of biological samples such as urine roids, including cortisol and cortisone, were quantitatively de-
and serum. However, the methods have still hampered from tectable (11).
incorrect physiological levels of adrenal steroids, including
cortisol, due to the sampling problems with circadian STRESS-MEDIATED HAIR CORTISOL IN ENDOCRINE
variation. In general, acute cortisol levels fluctuate markedly SYSTEM
depending on many physiological factors including circadian
rhythmicity, and it may provide a rather poor reflection of nor- Because more than 70% of diseases are believed to be
mal, chronic cortisol secretion (12, 13). stress-related, prediction of chronic stress is an important step
　 In contrast to the biological fluids, hair can provide bio- in reducing the incidence of chronic illnesses. Hair cortisol
logical information of long-term exposure because its growth may provide an objective measurement of stress over time
rate is about 1 cm/month (Fig. 3). The hormones are mainly rather than just ‘a day in life’. The symptoms of the metabolic
delivered from the blood circulation to the capillaries of the syndrome resemble those of Cushing’s syndrome, a disease
dermal papilla, which is located in the hair follicles (33). This that is characterized by hypercortisolism. One of the questions
phenomenon enables retrospective examination of cortisol raised is whether chronically elevated cortisol concentrations
production at the times when a stressor is most salient, without play a role in the development of obesity and the metabolic
needing to take a sample right at that time. Alternatively, it can syndromes. The increased hair cortisol levels are also asso-
provide a baseline cortisol assessment for a time period during ciated with children’s obesity caused by long-term activation
which the stress has not yet occurred. In addition, its non-in- of HPA-axis (39), which is in accordance with the urinary lev-
vasive nature in sampling and easy storage at room temper- els of cortisol in obese children (40). The risk of cardiovascular
ature has been highlighted as advantage in clinical applica- disease (CVD) is associated with the increase in hair cortisol
tions (34-36). In the hair analysis, steroid hormones are ex- levels by 2.7-times, which was similar to the risk associated with
tracted by solubilization or digestion of the hair matrix with al- hypertension or obesity. This suggests that high cortisol levels in
kaline hydrolysis for androgen and estrogens (37, 38). Also, a long-term might be an important risk factor for CVD (41).
the acid hydrolysis or methanol extraction can enable an anal- 　Mitotane, an anti-neoplastic agent for adrenocortical cancer
ysis of corticoids in hair due to its insufficient chemical stabil- (ACC), increases CBG and induces CYP3A4 activity, which
ity (11). When 62 biologically active steroids were analyzed leads to high doses of hydrocortisone; however, there has
by an optimized extraction technique, and only 20 hair ste- been no efficient biomarker to evaluate this therapy. Hair corti-
sol levels were higher in ACC patients compared to healthy in-
dividuals, and they were associated with body mass index
(42). However, there was no correlation between hair cortisol
levels and hydrocortisone doses. As a measure of long-term
cortisol exposure, the hair cortisol analysis in patients receiv-
ing glucocorticoid replacement therapy may be a useful tool.
Also, the hair cortisol content is correlated with hydro-
cortisone dose in the patients with adrenal insufficiency, who
had significantly higher subjective stress scores than the con-
trol subjects (43).

METABOLOMIC IDENTIFICATIONS IN CHRONIC

STRESS

The main benefit of metabolomic strategy is the high like-

lihood of identifying unpredicted changes in metabolic profiles
cued by abnormal conditions (44). In particular, the metab-
olomic information may offer novel diagnostic indicators and
therapeutic targets in clinical applications (45), as well as in
mechanistic studies heading to elucidate metabolic modules that
can regulate dysfunctional processes in disease statuses (46).
　Among dysfunctional heath conditions, chronic diseases are
mostly composed of subtle and long-term dysregulations of
Fig. 3. A proposed mechanism of cortisol incorporation into hair cellular and physiological function that is often not measurable
and retrospective reflection of its chronic secretion. Cortisol may
be incorporated into hair via passive diffusion from (A) blood ca-
even during disease onset (47). The stress-inducing abnormal
pillary, (B) sweat, and (C) sebum, as well as from (D) external status is chronically developed with unpredictable combina-
sources. Data are taken from reference #12. tion of various types of etiological factors. Therefore, the mo-

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 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

lecular characterization as well as clinical definition may not source caused by transient (time-course) or individual specific-
be clear at specific disease course and it may not be homolo- ity can be handled within reconstructed statistical model,
gous across individuals (48). The diversity of pathological traits which in turn offers more robust candidates for diagnostic/
induced by chronic stress make it even harder to properly di- prognostic information in clinical cases (Fig. 4).
agnose the abnormality and classify the progress stage. Most of 　In many studies, metabolomic approach has been limitedly
the studies are mainly focused on targeted single molecule and applied to biomarker discovery with either single target or
one-point interaction, which may be insufficient to reflect the group of metabolites. Yet, an ideal exploratory use of the tech-
dynamics and systematic effects of chronic stress (49). In this nology would not only be able to capture the metabolite
context, metabolite profiling can be an effective tool for bio- changes associated with pathology, but it would also elucidate
marker discovery and understanding of molecular mechanism, the molecular mechanism responsible for the dysfunction and
particularly for the cases where a diagnostic/prognostic in- propose the logical candidate for regulating the abnormality.
dicator is unknown and molecular mechanism remains veiled. Pathway (enrichment) analysis, a systematic approach for data
Recently, a chronic unpredictable mild stress in animal model mining from metabolite sets, can provide metabolic path-
showed aberrant profiling of amino acids that were grouped way-wise information rather than the readouts of individual or
into neurotransmitters and branched-chain amino acids (50). few metabolite contents (52). This bioinformatics tool, which
The metabolic signatures under acute and chronic stress in rat is rooted in gene ontology enrichment analysis, assigns groups
model and the biochemical cues are also closely associated of metabolites at the level of metabolic pathway. The analysis
with behavior and physiological readouts (49). provides P value that presents significant difference at the lev-
els of metabolic pathways and information on pathway cen-
DATA MINING IN METABOLOMICS trality (pathway importance), which demonstrates how much
connectivity is formed surrounding the metabolites of interest
To capture disease-specific metabolic signatures, many studies, in the pathway (53). The output can be further investigated for
including the case studies described above, primarily explored mechanistic understanding of disease, and it can be applied
on the multiple molecular constituents (metabolites), robustly for targeting and controlling the functional points of disease
managed variability and heterogeneity of disease progress, and process. However, although metabolic signature can be identi-
defined the individuality of metabolic contents, which could fied at the level of either single molecule, groups of metabo-
lead to well-defined diagnoses and prognoses. In this context, lites, or metabolic pathway, metabolite assessment cannot by
multivariate statistics such as principal component analysis itself provide a complete understanding of disease process. To
(PCA) or partial least squared algorithm is a useful data mining gain comprehensive causality and mechanism in given ex-
tool that can distinguish different groups with minimized loss perimental designs, integrative analysis should be accom-
of information (51). By this nature, unidentified variation panied by other molecular information such as mRNA ex-

Fig. 4. Reconstructed metabolic net-

work for systematic screening of ther-
apeutic target point. The re-modeled
metabolic structure is composed of
metabolites, which consists of node
(e.g. metabolite) and edge (e.g. corre-
lation, structural similarity). The model
may be extended to protein infor-
mation via reaction pair that has al-
ready been built in the assembly,
which could unravel “hidden” or veiled
metabolic modulation, particularly in
chronic disease. In this example, node
color and size indicate significant dif-
ference and fold change compared to
control (e.g. disease vs control), while
edge presents two layers of informa-
tion on structural similarity and re-
action likelihood, which leads to auto-
matic rearrangement as seen in the
figure indicating proximity of bioche-
mical module.

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 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

pression levels and protein abundances (54). Since metab- not only be restricted to cortisol (35). Further research on es-
olomics joins systems with biology, mRNA has been the first tablishing the reference values of hair cortisol may be needed
partner for integrative analysis mainly due to the well-estab- to enhance the current knowledge on particular aspects of
lishment of technical platform and advancements in statistical chronic stress and other metabolic changes in many endocrine
analysis. However, the expression level of mRNA showed a diseases.
low correlation with enzyme activity and protein expression
levels, which is a direct molecular companion with metabo- REFERENCES
lites, the substrate, and products. Thus protein information is
more suitable counterpart of metabolite readout (55). 1. Anderson NB (1998) Levels of analysis in health sciences:
　Most importantly, in order to have molecular information A framework for integrating sociobehavioral and bio-
(mRNA, protein, and metabolite) located at the cornerstone of medical research. Ann N Y Acad Sci 840, 563-576
clinical application, various types of molecular levels should 2. Edwards C (2012) Sixty tears after hench-Corticosteroids
and chronic inflammatory disease. J Clin Endocrinol
be linked to physiological traits (e.g. clinical parameter) in a
Metab 97, 1443-1451
statistically sound manner. This has been mainly done by 3. Whirledge S and Cidlowski A (2013) A role for glucocorti-
‘qualitative’ assessment in which the final readout of molec- coids in stress-impaired reproduction: Beyond the hypo-
ular behaviors is linked to logical phenotypes of disease. For thalamus and pituitary. Endocrinology 154, 4450-4468
example, aberrant glycolytic activity can be associated with tu- 4. Oswald LM, Zandi P, Nestadt G, Potash JB, Kalaydjian AE
mor metabolism, in which glycolytic intermediates and key and Wand GS (2006) Relationship between cortisol re-
enzymes are overexpressed in tumor cells. But in the case of sponse to stress and personality. Neuropsychopharmacology
chronic disease, we may not be able to detect “the stand- 31, 1583-1591
ardized feature” like cancer metabolism case. Alternatively, 5. Staab CA and Maser E (2010) 11-Hydroxysteroid de-
hydrogenase type 1 is an important regulator at the inter-
we can track and correlate the quantitative traits of pathology
face of obesity and inflammation. J Steroid Biochem Mol
with molecular dynamics, which can bear the variability Biol 119, 56-72
caused by individuality and differential disease progress. One 6. Juster RP, McEwen BS and Lupien SJ (2010) Allostatic load
relevant statistical approach is canonical correlation analysis biomarkers of chronic stress and impact on health and
(CCA), which has been often applied in psychological, cli- cognition. Neurosci Biobehav Rev 35, 2-16
mate, and ecological studies to enumerate the correlations be- 7. Miller WL (2008) Steroidogenic enzymes. Endocr Rev 13,
tween two distinct data sets measured on the identical ex- 1-18
perimental units (56-59). This statistics can analyze significant 8. Weitzman ED, Fukushima D, Nogeire C, Roffwarg H,
relations between two different dataset (e.g. metabolite con- Gallagher TF and Hellman L (1971) Twenty-four hour pat-
tern of the episodic secretion of cortisol in normal
tents and the associated clinical parameters). It is similar as
subjects. J Clin Endocrinol Metab 33, 14-22
PCA, in the way that CCA seeks for linear combinations of the 9. Hellhammer DH, Wüst S and Kudielka BM (2009)
variables to reduce the dimension of the data sets; but at the Salivary cortisol as a biomarker in stress research. Psycho-
same time, it explores to maximize the correlation between neuroendocrinology 34, 163-171
the two variates (58). 10. Elias PCL, Martinez EZ, Barone BFC, Mermejo LM, Castro
M and Moreira AC (2014) Late-night salivary cortisol has a
CONCLUDING REMARKS better performance than urinary free cortisol in the diag-
nosis of Cushing’s syndrome. J Clin Endocrinol Metab 99,
Because single cortisol assessments are strongly affected by the 2045-2051
11. Jung HJ, Kim SJ, Lee WY, Chung BC and Choi MH (2011)
acute context of the measurement situation (time of day, day
Gas chromatography/mass spectrometry based hair steroid
of the week, and other circumstances such as distress for profiling may reveal pathogenesis in hair follicles of the
blood sampling), the assessment of long-term cortisol secretion scalp. Rapid Commun Mass Spectrom 25, 1184-1192
from the biological fluids, such as blood, urine, and saliva, is 12. Stalder T and Kirschbaum C (2012) Analysis of cortisol in
highly labor-intensive and rather difficult to be implemented in hair - State of the art and future directions. Brain Behav
physical and psychological studies. Hair growth patterns also Immun 26, 1019-1029
vary across different regions of the scalp, and the precise 13. Russell E, Koren G, Rieder M and Uum SV (2012) Hair
mechanisms of substances incorporating into hair are still in- cortisol as a biological marker of chronic stress: Current
completely understood. But a growing observation supports status, future directions and unanswered questions. Psy-
choneuroendocrinology 37, 589-601
the notion that hair cortisol analysis provides a valid and reli-
14. Dimsdale JE and Moss J (1980) Short-term catecholamine
able reflection of long-term cortisol secretion (12, 13, 27). We response to psychological stress. Psychosom Med 42,
mainly discussed about the advantages of hair analysis as an 493-497
index of chronic cortisol response. Due to their association 15. Karatsoreos IN and McEwen BS (2011) Psychobiological
with various aspects of biochemical stress in adrenal steroids allostasis: resistance, resilience and vulnerability. Tr Cognit
and other steroid metabolites, advantages of hair analysis may Sci 15, 576-584

214 BMB Reports http://bmbreports.org

 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

16. McEwen BS (1998) Protective and damaging effects of 34. Choi MH, Kim KR, Kim IS, Lho DS and Chung BC (2001)
stress mediators. N Engl J Med 338, 171-179 Increased hair polyamine levels in patients with
17. Seeman TE, McEwen BS, Rowe JW and Singer BH (2001) Alzheimer’s disease. Ann Neurol 50, 128
Allostatic load as a marker of cumulative biological risk: 35. Cho SH, Choi MH, Sim WY, Lee WY and Chung BC
MacArthur studies of successful aging. Proc Natl Acad Sci (2010) Metabolic alterations of DHEA and cholesterol sul-
U S A 98, 4770-4775 phates in the hair of patients with acne measured by liq-
18. Staufenbiel SM, Penninx BW, Spijker AT, Elzinga BM and uid chromatography-mass spectrometry. Exp Dermatol 19,
van Rossum EF (2013) Hair cortisol, stress exposure, and 694-696
mental health in humans: a systematic review. Psychoneu- 36. Choi MH, Kim SJ, Lew BL, Sim WY and Chung BC (2013)
roendocrinology 38, 1220-1235 Hair steroid profiling reveals racial differences in male
19. Sapolsky RM, Krey LC and McEwen BS (1986) The pattern baldness between Korean and Caucasian popul-
Neuroendocrinology of Stress and Aging: The Glucocorti- ations. J Invest Dermatol 133, 822-824
coid Cascade Hypothesis. Endocr Rev 7, 284-301 37. Choi MH and Chung BC (1999) GC-MS determination of
20. Mizoguchi K, Ishige A, Aburada M and Tabira T (2003) steroids related to androgen biosynthesis in human hair
Chronic stress attenuates glucocorticoid negative feed- with pentafluorophenyldimethylsilyl-trimethylsilyl deriva-
back: involvement of the prefrontal cortex and hippo- tisation. Analyst 124, 1297-1300
campus. Neuroscience 119, 887-897 38. Choi MH, Kim KR and Chung BC (2000) Determination of
21. Hamrahian AH, Oseni TS and Arafah BM (2004) Measure- estrone and 17-estradiol in human hair by gas chroma-
ment of serum free cortisol in critically ill patients. N Engl tography-mass spectrometry. Analyst 125, 711-714
J Med 350, 1629-1638 39. Veldhorst MA, Noppe G, Jongejan MH et al (2014)
22. Levine A, Zagoory-Sharon O, Feldman R, Lewis JG and Increased scalp hair cortisol concentrations in obese
Weller A (2007) Measuring cortisol in human psycho- children. J Clin Endocrinol Metab 99, 285-290
biological studies. Physiol Behav 90, 43-53 40. Reinehr T, Kulle A, Wolters B et al (2014) Relationship be-
23. Yamada J, Stevens B, de Silva N et al (2007) Hair cortisol tween 24-hour urinary free cortisol concentrations and
as a potential biologic marker of chronic stress in hospi- metabolic syndrome in obese children. J Clin Endocrinol
talized neonates. Neonatology 92, 42-49 Metab 99, 2391-2399
24. Groeneveld MG, Vermeer HJ, Linting M, Noppe G, van 41. Manenschijn L, Schaap L, van Schoor NM et al (2013)
Rossum EF and van IJzendoorn MH (2013) Children's hair High long-term cortisol levels, measured in scalp hair, are
cortisol as a biomarker of stress at school entry. Stress 16, associated with a history of cardiovascular disease. J Clin
711-715 Endocrinol Metab 98, 2078-2083
25. Pereg D, Gow R and Mosseri M (2011) Hair cortisol and 42. Manenschijn L, Quinkler M and van Rossum EF (2014)
the risk for acute myocardial infarction in adult men. Hair cortisol measurement in mitotane-treated adrenocort-
Stress 14, 73-81 ical cancer patients. Horm Metab Res 46, 299-304
26. Pereg D, Chan J, Russell E et al (2013) Cortisol and testos- 43. Gow R1, Koren G, Rieder M and Van Uum S (2011) Hair
terone in hair as biological markers of systolic heart cortisol content in patients with adrenal insufficiency on
failure. Psychoneuroendocrinology 38, 2875-2882 hydrocortisone replacement therapy. Clin Endocrinol
27. Stalder T, Kirschbaum C, Alexander N et al (2013) Cortisol (Oxf) 74, 687-693
in hair and the metabolic syndrome. J Clin Endocrinol 44. Lee do Y, Kind T, Yoon YR, Fiehn O and Liu KH (2014)
Metab 98, 2573-2580 Comparative evaluation of extraction methods for simulta-
28. Steudte S, Kirschbaum C, Gao W et al (2013) Hair cortisol neous mass-spectrometric analysis of complex lipids and
as a biomarker of traumatization in healthy individuals primary metabolites from human blood plasma. Anal
and posttraumatic stress disorder patients. Biol Psychiatry Bioanal Chem 406, 7275-7286
74, 639-646 45. Crews B, Wikoff WR, Patti GJ et al (2009) Variability anal-
29. Vining RF, McGinley RA and Symons RG (1983) Hormones ysis of human plasma and cerebral spinal fluid reveals
in saliva: mode of entry and consequent implications for statistical significance of changes in mass spectrome-
clinical interpretation. Clin Chem 29, 1752-1756 try-based metabolomics data. Anal Chem 81, 8538-8544
30. Cho HJ, Kim JD, Lee WY, Chung BC and Choi MH (2009) 46. Vinayavekhin N, Homan EA and Saghatelian A (2009)
Quantitative metabolic profiling of 21 endogenous corti- Exploring disease through metabolomics. ACS Chem Biol
costeroids in urine by liquid chromatography-triple quad- 5, 91-103
rupole-mass spectrometry. Anal Chim Acta 632, 101-108 47. Kim OY, Lee JH and Sweeney G (2013) Metabolomic
31. Moon JY, Jung HJ, Moon MH, Chung BC and Choi MH profiling as a useful tool for diagnosis and treatment of
(2009) Heat-map visualization of gas chromatography- chronic disease: focus on obesity, diabetes and cardio-
mass spectrometry based quantitative signatures on ste- vascular diseases. Expert Rev Cardiovasc Ther 11, 61-68
roid metabolism. J Am Soc Mass Spectrom 20, 1626-1637 48. Li ZY, Zheng XY, Gao XX et al (2010) Study of plasma
32. Maier B and Vogeser M (2013) Target analyte quantifica- metabolic profiling and biomarkers of chronic un-
tion by isotope dilution LC-MS/MS directly referring to in- predictable mild stress rats based on gas chromatog-
ternal standard concentrations-validation for serum corti- raphy/mass spectrometry. Rapid Commun Mass Spectrom
sol measurement. Clin Chem Lab Med 51, 833-837 24, 3539-3546
33. Randall VA (1994) Androgens and human hair growth. 49. Wang X, Zhao T, Qiu Y et al (2009) Metabonomics ap-
Clin Endocrinol (Oxf) 40, 439-457. proach to understanding acute and chronic stress in rat

http://bmbreports.org BMB Reports 215

 Metabolic biomarkers in chronic stress
Do Yup Lee, et al.

models. J Proteome Res 8, 2511-2518 J and Weckwerth W (2008) Integration of metabolomic

50. Ni Y, Su M, Lin J et al (2008) Metabolic profiling reveals and proteomic phenotypes analysis of data covariance dis-
disorder of amino acid metabolism in four brain regions sects starch and RFO metabolism from low and high tem-
from a rat model of chronic unpredictable mild stress. perature compensation response in arabidopsis thaliana.
FEBS Lett 582, 2627-2636 Mol Cell Proteomics 7, 1725-1736
51. Broadhurst DI and Kell DB (2006) Statistical strategies for 56. Meyer RC, Steinfath M, Lisec J et al (2007) The metabolic
avoiding false discoveries in metabolomics and related signature related to high plant growth rate in Arabidopsis
experiments. Metabolomics 2, 171-196 thaliana. Proc Natl Acad Sci U S A 104, 4759-4764
52. Xia J and Wishart DS (2010) MetPA: a web-based metab- 57. Jozefczuk S, Klie S, Catchpole G et al (2010) Metabolomic
olomics tool for pathway analysis and visualization. and transcriptomic stress response of Escherichia coli. Mol
Bioinformatics 26, 2342-2344 Syst Biol 6, 364
53. Xia J, Mandal R, Sinelnikov IV, Broadhurst D and Wishart 58. González I, Déjean S, Martin PG, Gonçalves O, Besse P
DS (2012) MetaboAnalyst 2.0-a comprehensive server for and Baccini A (2009) Highlighting relationships between
metabolomic data analysis. Nucleic Acids Res 40, W127- heterogeneous biological data through graphical displays
W133 based on regularized canonical correlation analysis. J Biol
54. Lee do Y, Park JJ, Barupal DK and Fiehn O (2012) System Syst 17, 173-199
response of metabolic networks in Chlamydomonas rein- 59. Song IS, Lee do Y, Shin MH et al (2012) Pharmacogene-
hardtii to total available ammonium. Mol Cell Proteomics tics meets metabolomics: discovery of tryptophan as a
11, 973-988 new endogenous oct2 substrate related to metformin
55. Wienkoop S, Morgenthal K, Wolschin F, Scholz M, Selbig disposition. PLoS One 7, e36637

216 BMB Reports http://bmbreports.org