KR101029251B1 - Evaluation Method and Evaluation Indicator of Cortisol Diurnal Rhythm by Determination of Salivary Cortisol - Google Patents

Abstract

The present invention relates to an index for determining the normality of cortisol circadian rhythm by measuring cortisol concentration contained in saliva and cortisol circadian rhythm evaluation method based on the concentration of cortisol in saliva. Vapor cortisol concentrations, CAR cortisol concentrations and bedtime cortisol concentrations, which are the cortisol concentrations in saliva immediately after waking up, 30 minutes after waking up and just before bedtime, were measured. One or more of an increase in CAR cortisol concentration based on concentration, a decrease in bed cortisol concentration based on CAR cortisol concentration, a percentage increase in CAR cortisol concentration based on gaseous cortisol concentration, and a percentage decrease in bed cortisol concentration based on CAR cortisol concentration The present invention relates to a method for evaluating normal rhythm of cortisol using selected factors.

Cortisol, adrenal glands, saliva, cortisol awakening response, hormones, circadian rhythms

Description

Evaluation Method and Evaluation Indicator of Cortisol Diurnal Rhythm by Determination of Salivary Cortisol}

The present invention can quickly and objectively determine whether cortisol circadian rhythm is normal by measuring cortisol concentration contained in saliva, and cortisol circadian rhythm evaluation indicators for determining whether adrenal function and hormone secretion function are normal and It is about an evaluation method.

The adrenal gland is located in the shape of a cone above the kidneys. The adrenal cortex is histologically divided into glomerulosa, fasciculata and redicularis. Cortisol is mainly produced from the fasciculata, and dehydroisoandrosterone is produced mainly from the redicularis. do.

The hormone production of the adrenal glands has a very elaborate circadian rhythm (or Diurnal Rhythm) and is the only endocrine organ in the body that produces hormones in response to external and external stimuli (stresses). A patent has been applied for measuring and evaluating an individual's stress index by using a concentration of corticosteroids using this feature (WO 2002/62198).

The daily rhythm of the adrenal glands is known to be controlled by the cranial nerves, hypothalamus, and pituitary and is essentially determined by the individual sleep cycles. After a normal night's sleep, waking up in the morning, the brain's nerves wake up, and this signal reaches the hypothalamus of the suprachiasmatic nucleus (SCN), and in the intersecting nucleus, the bioclock gene (bioclock). The gene is activated and this signal is transmitted back to the paraventricular nucleus (PVN). In the ventricular nucleus, corticotropin releasing hormone (CRH) is secreted in response to this stimulus, and the secreted secretory hormone stimulates the pituitary gland to produce and secrete adrenocorticotrophic hormone (ACTH). . The levels of corticosteroids produced and secreted by the pituitary gland peak at 15 minutes after waking up. Corticosteroids secreted in the blood act on the adrenal glands and increase the production of cortisol in the adrenal cortex. The peak cortisol concentration is about 30 minutes after waking up, with the highest cortisol concentration of the day. As such, a phenomenon in which cortisol concentration reaches a normal level in about 30 minutes after waking up in the morning is called a cortisol awakening response (hereinafter referred to as CAR).

CAR is not affected by the general sleeping environment, light, the phenomenon of waking during sleep time, and is known to be affected by health conditions. Patients with systemic diseases such as diabetes, cardiovascular disease, and cancer are known to show dull CAR with low concentration difference between cortisol released immediately after waking up and 30 minutes after waking up. Depression, fatigue, migraine, post-traumatic syndrome Dull CARs are known to appear in patients with disease, and dull CARs are known to appear in healthy but older people. However, factors such as stress, anxiety, and worry from daily work are reported to play a role in boosting CAR.

Taken together, these studies show that the increase in cortisol concentration after morning weather indicates the overall homeostatic state of the body and acts as a biomarker of the adrenal glands that collectively shows changes in mind and body due to stress. So far, about 160 papers have published the results of research on systemic diseases and mental and physical diseases using CAR as a biomarker. The relationship between CAR reported so far and systemic and mental and mental diseases is summarized in Table 1 and Table 2.

Table 1 Diseases Represented by Slowing CAR

Table 2 Diseases Increasing CAR

In the meantime, hormone concentrations were measured mainly in blood. From an endocrinology point of view, blood is not a hormone's target organ, but a simple hormone carrier. Most blood protein hormones (growth hormones, insulin, etc.) are well soluble in water and are present in activated form in the blood, but steroids produced by the adrenal cortex and gonads are insoluble in water, so they can be combined with other media (carrier proteins). Is present in the blood. When steroids are combined with the carrier, they do not act on the hormone receptors in the target organs, so they are called "inactive states" and cannot induce physiological mechanisms. Most of the steroids in the blood (95-99%) are present in this inactive state and only a fraction (1-4%) are known to be in an unbound state (active state). Therefore, the concentration of steroids in the blood can be said to be the concentration of most inactive rather than the concentration of the hormone in the active state.

Unlike blood, all of the steroids in saliva are not active with the carrier medium, and active steroids contained in the extracelluar fluid are transferred to the salivary glands by passive diffusion. will be. Therefore, the steroid concentration contained in saliva is recognized as an active steroid contained in the tissue.

In addition, saliva, unlike blood, can be easily collected by the general public in accordance with the individual's sleep cycle without the help of medical personnel, so it has the advantage of easily conducting research on the hormonal circadian rhythm of the individual.

In other words, using saliva has the advantage of easy access to the concentration of active steroids and the hormonal circadian rhythms, which were difficult to access to blood. However, because the concentration of steroids in saliva is as low as 1 / 100-1 / 1000 of the blood, it cannot be analyzed by commercially available hormone analysis kits.

CARs are closely related to an individual's sleep cycle, so measuring cortisol levels in the blood is virtually impossible unless hospitalized. For this reason, individual CARs have been mainly determined by measuring the concentration of cortisol contained in saliva. CAR is not simply a response to the adrenal gastrointestinal tract, but a response of the hypothalamus-pituitary-adrenal axis (HPA axis) according to the weather, which is a brain of the hormone production and regulation of the hypothalamus. You can guess the rhythm.

Since the change in the cyclic rhythm of the hormone is linked to the physiological rhythm of the whole body, it is known that the CAR according to the weather is connected to the increase in blood glucose, body temperature, pulse rate, and metabolism required after the weather.

As described above, although many studies have been conducted by using the CAR which appears after the weather as a biomarker in various research fields, comparative comparison / analysis is mostly performed on a case-by-case basis, and there are no quantified and objective criteria for evaluating the CAR. .

An object of the present invention is to provide an indicator and an evaluation method for determining the normal secretion rhythm of the hormone, in detail, an evaluation indicator for determining whether the cortisol circadian rhythm is normal and the evaluation to determine whether the cortisol circadian rhythm is normal It provides a method, and provides an indicator and method that can quickly, accurately and objectively determine whether the cortisol circadian rhythm is normal, and an indicator that can determine whether the adrenal function and hormone secretion function normal using cortisol To provide a way.

The indicator according to the present invention is an indicator for determining whether the hormone secretion function is normal and whether the adrenal function is normal, and is an indicator for determining whether the hormone secretion and adrenal function is normal using saliva cortisol. Characteristically, the cortisol circadian rhythm is determined to determine whether normal hormone secretion and adrenal function are normal.

In detail, the cortisol circadian rhythm evaluation index according to the present invention is an index for determining whether cortisol circadian rhythm is normal and at the same time an indicator for determining whether adrenal function and hormone secretion function are normal. 1) meteorological indicators of cortisol concentrations in saliva immediately after the weather; 2) CAR indicators of the saliva cortisol concentration in the cortisol reactor according to the gas phase; 3) sleep indicators, the concentration of cortisol in saliva just before bedtime; 4) CAR growth index, which is an increase in the saliva cortisol concentration of the cortisol reactor according to the gas phase based on the saliva cortisol concentration immediately after the gas phase; 5) CAR reduction index which is the amount of reduction in the saliva cortisol concentration immediately before bed based on the saliva cortisol concentration in the cortisol reactor according to the gas phase; 6) CAR growth rate indicator which is an increase percentage of saliva cortisol concentration in the cortisol reactor according to the gas phase based on the saliva cortisol concentration immediately after the gas phase; And 7) a CAR reduction rate indicator, which is a percentage reduction of the saliva cortisol concentration immediately before bed based on the cortisol concentration in the saliva of the cortisol reactor according to the gas phase.

1 is a cortisol circadian rhythm evaluation indicators according to the present invention on the basis of changes in daily cortisol concentration by weather and bedtime (1) weather indicator, 2) CAR indicator, 3) sleeping indicator, 4) CAR increase index, 5) CAR decrease Indicators, 6) CAR growth rate indicators, and 7) CAR decrease rate indicators.

The cortisol reactor according to the gas phase means a cortisol awakening response (CAR) where the cortisol concentration reaches the highest level of the day in about 30 minutes after waking up in the morning, and substantially means 30 minutes after the gas phase. do.

The indicator is an indicator for determining cortisol circadian rhythm as normal, the weather indicator is 3.37 ~ 16.62 nmol / L, the CAR indicator is 6.83 ~ 31.47 nmol / L, the sleep indicator is 0.73 ~ 5.10 nmol / L , The CAR growth index is 1.85 ~ 17.33 nmol / L, the CAR reduction index is 4.60 ~ 26.65 nmol / L, the CAR growth index is 22.64 ~ 214.67%, the CAR reduction index is 63.10 ~ 94.09% have.

The evaluation method according to the present invention is a method for determining whether the hormone secretion function is normal and whether the adrenal function is normal, characterized in that it is a method for determining whether the hormone secretion and adrenal function is normal by using saliva cortisol concentration, More specifically, it is a method of determining whether the cortisol circadian rhythm is normal and whether the hormone secretion and adrenal function are normal in the body.

In detail, the cortisol circadian rhythm evaluation method according to the present invention is evaluated using the concentration of cortisol in saliva, the cortisol concentration in the saliva immediately after the weather, 30 minutes after the weather and just before bedtime cortisol (cortisol) concentration Concentration, CAR cortisol concentration and bedtime cortisol concentration were measured, the amount of increase of CAR cortisol concentration based on gaseous cortisol concentration, the amount of decrease of bed cortisol concentration based on CAR cortisol concentration, the percentage of increase of CAR cortisol concentration based on gaseous cortisol concentration, and It is characterized by determining whether cortisol diurnal rhythm is normal using one or more selected factors from the percentage of reduction in bedtime cortisol concentration based on the CAR cortisol concentration.

The saliva cortisol (cortisol) concentration is characterized by radioimmunoassay, the factor is the gaseous cortisol concentration, CAR cortisol concentration, bedtime cortisol concentration, the increase in the amount of CAR cortisol concentration based on the gas phase cortisol concentration, CAR cortisol concentration It is characterized by the amount of reduction in the standard bedtime cortisol concentration, the percentage of increase in the CAR cortisol concentration based on the vapor phase cortisol concentration, and the percentage of decrease in the bedtime cortisol concentration based on the CAR cortisol concentration.

Characteristically, the evaluation method according to the present invention is the gas phase cortisol concentration is 3.37 ~ 16.62nmol / L, the CAR cortisol concentration is 6.83 ~ 31.47nmol / L, the bedtime cortisol concentration is 0.73 ~ 5.10nmol / L, gaseous phase The increase in the amount of CAR cortisol based on the cortisol concentration is 1.85 to 17.33 nmol / L, and the decrease in bedtime cortisol concentration based on the CAR cortisol concentration is 4.60 to 26.65 nmol / L, and the increase in the concentration of CAR cortisol based on the gas phase cortisol concentration If the percentage is 22.64 to 214.67% and the percentage of decrease in bedtime cortisol concentration based on the CAR cortisol concentration is 63.10 to 94.09%, the cortisol circadian rhythm is determined to be normal, and if the cortisol circadian rhythm is determined to be normal, the body Hormonal secretion and adrenal function are distinguished by normal.

Indicators of the present invention can determine the normal status of hormone secretion and adrenal function in the body, as well as cortisol circadian rhythms, and cortisol circumference as assessed by relative comparison, clinical practice, or any variable criteria Rhythms and hormonal secretions in the body can be accurately, objectively and easily evaluated, and there is an advantage in determining whether or not a disease related to cortisol secretion or a disease causing stress is present.

The evaluation method of the present invention can objectively and quickly determine whether cortisol circulatory rhythm is normal, and whether adrenal function is normal or not, and it is more accurate and reliable by using an active cortical hormone. High evaluation can be performed, and there is an advantage that it is possible to comprehensively and accurately assess the function of hormones and organs related to hormone secretion.

Hereinafter, the indicators and the evaluation method of the present invention will be described in detail with reference to the accompanying drawings. The drawings introduced below are provided by way of example so that the spirit of the invention to those skilled in the art can fully convey.

Hereinafter, the technical and scientific terms used herein will be understood by those skilled in the art without departing from the scope of the present invention. Descriptions of known functions and configurations that may be unnecessarily blurred are omitted.

Screening of Indicators and Extracted Indicators

Saliva was collected from 133 men and women (12-63 years) with no disease for two days a year, immediately after waking up, 30 minutes after waking up, and immediately before bedtime. Saliva was collected using a saliva collection tube, and food, tea, coffee smoking, and carbonated water were not consumed before saliva collection. Saliva cortisol concentrations were independent of flow rate, and sugar-free gums did not affect hormonal measurements, which allowed chewing sugar-free gums when the subject needed saliva stimulation. Before saliva sample collection, they were asked to rinse their mouths with water and to record the time of collection on the label provided.

The saliva samples collected were measured for saliva cortisol concentration using radioimmunoassay, in particular, liquid phased-double antibody method.

Pure cortisol used as a standard sample was purchased from Sigma-Aldrich. This standard cortisol was dissolved in an organic solvent in which ethanol and propylene glycol were mixed in a 1: 1 ratio, and phosphorylation buffer solution (GPBS, pH = 7.2) in which 0.1% gelatin was dissolved at a predetermined concentration. , Further diluted with GPBS) to prepare standard samples.

Cortisol antibodies were purchased from Biogenesis. Cortisol antibodies are 0.001% for aldosterone, 4.1% for 11-deoxycorticosterone, 5.7% for deoxycortisol, 0.5% for 21-deoxycortisol, and corti. It cross-reacted to 1.2% for corticosterone and less than 0.001% for other steroid hormones.

The procedure is briefly described, and for a more detailed measurement, see the applicant's paper on hormonal determination using liquid phased-double antibody methods (Salivary cortisol and DHEA levels in the Korean population: age- related differences, diurnal rhythm, and correlations with serum levels.Yonsei Med J. 2007; 48: 379-88).

Test tubes containing standard samples and 0.1 ml of samples in the test tubes, each containing GPBS instead of antibodies, were added to the test tube without cortisol to determine the specific reaction. Added charcoal-stripped saliva with activated carbon.

The total radiation dose of the ¹²⁵ I-labeled cortisol was approximately 20,000 cpm / tube and the antibody was diluted to bind 30-35%. The total volume of the first reaction was 0.4 ml per tube, and the standard and saliva samples were reacted at 4 ° C for 24 hours. The next day, the reaction was further reacted for 30 minutes at room temperature for the secondary reaction, and then 0.1 ml of the secondary antibody was added to each test tube and reacted for 30 minutes. Then, 0.5 ml of 10% PEG was added and reacted for 15 minutes and centrifuged at 1500 g for 20 minutes at 4 ° C. The supernatant was removed using a vacuum pump, and the radiation dose of the precipitate remaining in each test tube was measured using a gamma ray meter (Cobra 5005, PerkinElmer Life and Analytical Sciences). Each measured cortisol level was calculated from a standard curve using RIASMART software (PerkinElmer Life and Analytical Sciences).

Weather Indicator, CAR Indicator and Sleep Indicator

Immediately after the weather, ANOVA was performed on the concentration of cortisol before bedtime 30 and before bedtime, which are summarized in Table 3. The highest value is 30 minutes after waking up and the lowest value just before bedtime (F-value = 266.28, p-value <0.0001).

Table 3 Average Cortisol Concentration

Cortisol concentrations ranged from 1.09 nmol / L to 23.22 nmol / L immediately after vapor phase. Cortisol concentrations from 30 minutes after vapor phase ranged from 3.82 nmol / L to 42.17 nmol / L, and cortisol concentration before bedtime was 0.58 nmol / L. To 7.29 nmol / L. Table 4 summarizes the main percentiles of cortisol concentration for each time period.

Table 4 Percentiles of Cortisol Concentration (nmol / L) by Time

CAR growth index and CAR growth index

The difference between the cortisol concentration immediately after the weather and the 30 minute cortisol concentration after the weather (CARinc = 30 minute cortisol concentration after the vapor phase by sample-the cortisol concentration immediately after the weather, hereinafter called CARinc) was calculated. Figure is shown. CARinc means the absolute increase in cortisol 30 minutes after waking up immediately after waking up. The mean and standard deviation of CARinc were 7.97 nmol / L and 5.04 nmol / L, respectively. The median was 7.15 nmol / L and the interquartile range (IQR) was 3.46 ~ 10.93 nmol / L.

Since the cortisol concentrations vary from individual to 30 minutes after waking and after waking, the absolute value of cortisol increased from just after waking to 30 minutes is different.

Accordingly, the rate of increase of the 30 minute cortisol concentration after the vapor phase with respect to the absolute concentration of the cortisol concentration after the vapor phase (CARinc% = (30 minute cortisol concentration after the vapor phase by sample-cortisol concentration after the vapor phase) / cortisol concentration after the vapor phase ㅧ 100, unit%, or less) , CARinc%) were used to derive evaluation factors that were not sensitive to individual body characteristics. The CARinc% statistic refers to the relative increase in cortisol 30 minutes after wake up. The mean and standard deviation of CARinc% were 103.50% and 66.29%, respectively, with median 89.58% and interquartile range (IQR) of 52.90 ~ 143.46%.

Figure 3 is a diagram showing the CARinc% (%) as a histogram, summarized in Table 5 the calculated results of CARinc, CARinc%.

(Table 5)

CAR Reduction Index and CAR Reduction Rate Indicator

The difference between the 30-minute cortisol concentration and the pre-sleep cortisol concentration (CARdec = 30-minute cortisol concentration after wake-up for each sample-the cortisol concentration immediately before bed, hereinafter CARdec), and FIG. 4 shows the calculated CARdec (nmol / L) as a histogram. One drawing. CARdec is an absolute decrease in the concentration of cortisol before bed after 30 minutes of gas phase. The mean and standard deviation of CARdec were 14.37 nmol / L and 7.09 nmol / L, respectively. The median was 12.99 nmol / L and the interquartile range (IQR) was 9.05 ~ 19.39 nmol / L.

Because the concentration of cortisol after 30 minutes before bedtime and before bedtime varies between individuals, the absolute value of the reduced cortisol from 30 hours before bedtime to before bedtime appears different. Accordingly, the rate of decrease of cortisol concentration before going to bed for 30 minutes cortisol concentration after absolute gas phase (CARdec% = (30 minutes cortisol concentration after vapor phase by sample-cortisol concentration before bedtime) / 30 minutes cortisol concentration after vapor phase by sample ㅧ 100, Unit%, hereinafter, CARdec%) was used to derive evaluation factors that are not sensitive to individual body characteristics. The CARdec% statistic represents the relative decrease in cortisol just before bedtime for 30 minutes after waking up. The mean and standard deviation of CARdec% were 83.90% and 9.59%, respectively, with median 86.05% and interquartile range (IQR) of 80.58-90.43%.

5 is a diagram showing the calculated CARdec% as a histogram, and summarized the calculated results of CARdec and CARdec% in Table 6.

Table 6

Based on the above results, the cortisol circadian rhythm is normal, that is, the hormone secretion is normal and the adrenal function is normal. 2) CAR indicator of the saliva cortisol concentration of the cortisol reactor according to the gas phase, 3) Sleep indicator of the cortisol concentration in the saliva immediately before bedtime, 4) Saliva in the saliva of the cortisol reactor according to the gas phase based on the concentration of cortisol in the saliva immediately after the bed phase CAR increase index as an increase in cortisol concentration, 5) CAR decrease index as a decrease in saliva cortisol concentration just before bedtime based on cortisol concentration in the cortisol reactor according to the weather, 6) Cortisol concentration in saliva immediately after the weather CAR growth rate indicator, which is an increase percentage of saliva cortisol concentration in the cortisol reactor according to the gaseous phase, 7) Bedtime based on the cortisol concentration in the saliva of the cortisol reactor according to the gas phase CAR reduction rate index, which is a percentage reduction in saliva cortisol concentration, was extracted, 1) 3.37 ~ 16.62nmol / L weather index, 2) 6.83 ~ 31.47nmol / L CAR index, and 3) 0.73 ~ 5.10nmol / L bedtime. 4) 1.85 to 17.33 nmol / L CAR growth index, 5) 4.60 to 26.65 nmol / L CAR reduction index, 6) 22.64 to 214.67% CAR growth index, 7) 63.10 to 94.09% CAR reduction index Screened.

Blind test using extracted indicators

Cortisol concentration in saliva was measured in a similar manner to the screening of the above indicators, blind tests were performed using the extracted indicators, and the results of one test out of 30 or more tests are summarized in Tables 7 to 10.

Table 7 Test Results of Sample A

Table 8 Test Results of Sample B

Table 9 Test Results of Sample C

Abnormally assessed sample A in Table 7 was a sample of an artery ruptured brain surgery patient included in Willis's disease, and an abnormally evaluated sample B in Table 8 is a cancer patient undergoing chemotherapy (after chemotherapy 1). Weekly), and sample C, which was evaluated as normal in Table 9, was a sample of a normal person without disease as a result of a medical examination.

As can be seen from the test results, the indicators of the present invention can be used to determine whether cortisol circadian rhythms are normal and reliable, as well as to improve abnormality of corticosteroids based on the cortisol circadian rhythm evaluation index. In addition, abnormal reduction can be assessed to infer the presence or absence of diseases in Table 1 or Table 2 that may occur when corticosteroid abnormalities, and cortisol circadian rhythm evaluation methods affect the hormone secretion state in the body or adrenal function The presence or absence of a disease that affects can also be determined.

For example, in the case of brain surgery performed by rupture of an artery in Willis's disease, it can be seen that high cortisol concentrations may be accompanied by depression, anxiety, and sleep disorders.

For example, in cancer patients undergoing chemotherapy, chemotherapy affects cortisol production and production rhythms. Symptoms such as adrenal insufficiency (fatigue, lethargy, anorexia, sleep disorders, etc.) It can be seen that it can be accompanied in cancer patients.

Furthermore, it can be seen that the presence or absence of a specific lesion can also be determined by cortisol circadian rhythm measurement result by using the trend of cortisol circadian rhythm appearing for each lesion based on the cortisol circadian rhythm discrimination index according to the present invention. Can be.

For example, cortisol circadian rhythm measurements of a group of patients (16 patients) who underwent brain surgery due to rupture of an artery in Willis's disease showed that cortisol circadian rhythm was abnormal in the case of Willis's circulatory artery rupture. In particular, it indicates the abnormality due to high cortisol concentration on the basis of the sleep indicator, CAR increase indicator, CAR reduction indicator, CAR growth rate indicator, CAR reduction rate indicator all can confirm the tendency to show an abnormality with a-value.

1 is a diagram showing a cortisol circadian rhythm evaluation index according to the present invention based on the concentration of cortisol by bedtime to bedtime,

FIG. 2 is a diagram showing the difference between the cortisol concentration immediately after the vapor phase and the 30 minute cortisol concentration after the vapor phase in a histogram.

FIG. 3 is a graph showing the increase rate of the cortisol concentration in 30 minutes after the weather with respect to the cortisol concentration immediately after the gas phase in a histogram.

FIG. 4 is a histogram showing the difference in the concentration of cortisol immediately before bed to the concentration of cortisol 30 minutes after waking up.

FIG. 5 is a histogram showing the rate of decrease of cortisol concentration immediately before bed to 30 minutes cortisol concentration after gas phase.

Claims (7)

Cortisol Diurnal rhythm is an indicator for determining the normality, the indicator is a weather indicator which is the cortisol concentration in the saliva immediately after the weather; CAR indicator of the saliva cortisol concentration in the cortisol reactor according to the gas phase; Sleep indicator, which is the concentration of cortisol in saliva just before bedtime; CAR growth index, which is an increase in the saliva cortisol concentration of the cortisol reactor according to the gas phase based on the saliva cortisol concentration immediately after the gas phase; CAR reduction index which is a decrease amount of saliva cortisol concentration immediately before bed based on the saliva cortisol concentration of the cortisol reactor according to the gas phase; CAR growth rate indicator which is an increase percentage of the saliva cortisol concentration of the cortisol reactor according to the gas phase based on the saliva cortisol concentration immediately after the gas phase; Cortisol circadian rhythm evaluation index comprising a; rate of reduction of the saliva cortisol in the saliva immediately before bed based on the cortisol concentration in the saliva cortisol reactor according to the gas phase. The method of claim 1, The indicator is to determine the cortisol circadian rhythm as normal, the weather indicator is 3.37 ~ 16.62nmol / L, the CAR indicator is 6.83 ~ 31.47nmol / L, the sleep indicator is 0.73 ~ 5.10nmol / L , The CAR growth index is 1.85 ~ 17.33nmol / L, the CAR reduction index is 4.60 ~ 26.65nmol / L, the CAR growth index is 22.64 ~ 214.67%, the CAR reduction index is 63.10 ~ 94.09% Cortisol circadian rhythm evaluation index. Cortisol concentration, CAR cortisol concentration, and bedtime cortisol concentration, which are the cortisol concentrations in saliva immediately after waking up, 30 minutes after waking up, and before bedtime, were measured, and the amount of increase of CAR cortisol concentration based on the gastric cortisol concentration was measured. Cortisol circadian rhythm using one or more selected factors from a decrease in bedtime cortisol concentration based on CAR cortisol concentration, a percentage increase in CAR cortisol concentration based on gaseous cortisol concentration, and a percentage decrease in bed cortisol concentration based on CAR cortisol concentration A method of evaluating cortisol circadian rhythm using saliva cortisol to determine whether normal (diurnal rhythm) is normal. The method of claim 3, wherein The saliva cortisol (cortisol) concentration was measured by radioimmunoassay characterized in that the cortisol circadian rhythm evaluation method using the saliva cortisol. The method of claim 3, wherein The factors include gaseous cortisol concentration, CAR cortisol concentration, bedtime cortisol concentration, increase in CAR cortisol concentration based on gaseous cortisol concentration, decrease in bedtime cortisol concentration based on CAR cortisol concentration, and increase in CAR cortisol concentration based on gaseous cortisol concentration. A method for evaluating cortisol circadian rhythm using saliva cortisol, characterized in that it is a percentage reduction in bedtime cortisol concentration based on the percentage and CAR cortisol concentration. The method of claim 5, The evaluation method is The gas phase cortisol concentration is 3.37 ~ 16.62nmol / L, The CAR cortisol concentration is 6.83 ~ 31.47 nmol / L, The bedtime cortisol concentration is 0.73-5.10nmol / L, The increase in CAR cortisol concentration based on the gas phase cortisol concentration is 1.85 to 17.33 nmol / L, The amount of reduction of bedtime cortisol concentration based on the CAR cortisol concentration is 4.60 to 26.65 nmol / L, The percentage increase in CAR cortisol concentration based on gaseous cortisol concentration is 22.64-214.67%, If the percent reduction in bedtime cortisol concentration based on the CAR cortisol concentration is 63.10-94.09%, Cortisol circadian rhythm evaluation method using the saliva cortisol, characterized in that the normal determination of cortisol circadian rhythm. The method of claim 6, The evaluation method is When cortisol circadian rhythm is determined to be normal, cortisol circadian rhythm evaluation method using saliva cortisol characterized in that the normal determination of hormone secretion and adrenal function in the body.

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