Heartfulness meditation alters neuroendocrine profiles: A randomized controlled trial on hormones of stress and well-being

Abstract

Background:

Chronic stress disrupts the neuroendocrine system, leading to imbalances in neurotransmitters and stress hormones such as oxytocin, β endorphins and cortisol, contributing to mood disorders and poor emotional regulation. Complementary and alternative practices like meditation have shown promising results in stress regulation and mood elevation. Heartfulness (HFN) meditation, rooted in yogic traditions and incorporating yogic transmission, is an emerging technique to improve emotional resilience and hormonal homeostasis. Therefore, this study aimed to evaluate the effects of HFN meditation on oxytocin, β-endorphins, and cortisol. It also evaluated the changes in meditation depth using validated psychometric tools.

Methods:

A randomized controlled trial was conducted. Participants were divided into experimental and control groups. The experimental group practiced guided HFN meditation for 30 days. Following this, a crossover design was implemented in which, the control group participants were now given the intervention of HFN meditation. Biochemical markers (serum oxytocin, β-endorphins, cortisol) were measured at day 30, and day 60. Psychometric assessments included the Meditation Depth Questionnaire and the Positive and Negative Affect Schedule (PANAS).

Results:

HFN meditation significantly improved meditation depth (ΔM = –14.87, 95% CI [–23.61,–6.13], P = .001, r = 0.333) and positive affect (ΔM = –8.48, 95% CI [–12.03,–4.93], P < .001, r = 0.29), while reducing negative affect (ΔM = 7.70, 95% CI [3.81, 11.60], P < .001, r = 0.21). Oxytocin and endorphin levels increased (oxytocin ΔM = +88.18, P = .003, r = 0.355 and endorphin ΔM = +94.83, P = .003, r = 0.357), and cortisol decreased (ΔM = –133.55, P < .001, r = 0.661). After crossover, the control group exhibited similar improvements. Negative correlations were found between cortisol and both oxytocin and β-endorphins.

Conclusion:

HFN meditation significantly modulates stress-related neuroendocrine markers and enhances positive emotional states. By increasing the levels of these happy hormones and reducing cortisol, HFN presents a promising non-pharmacological intervention for improving mental health and stress resilience.

1. Introduction

Mental health has become a global public health concern, with rising rates of anxiety, depression and other stress-related disorders impacting the population globally.^[1] Stress impacts people of all ages but to varying extents at different stages in life. Young adults and teenagers experience academic pressure, social issues, and identity formation difficulties, resulting in high levels of stress and vulnerability to depression and anxiety.^[2] Middle-aged individuals often go through stress related to work pressure, financial problems, and family caregiving, leading to burnout and mental fatigue.^[3] At the same time, older people can face stressors like chronic disease, loneliness, and bereavement, which can worsen emotional distress and cognitive impairment.^[4]

Chronic stress can trigger a cascade of hormonal reactions in the body, primarily under the control of the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system.^[5] Dysregulation of these key hormones due to chronic stress contributes to a variety of health disorders, such as anxiety, depression, metabolic syndrome, and immune dysfunction.^[6] One of the most important hormones that mediate the stress response is cortisol, secreted by the adrenal glands when the HPA axis is activated. Although acute release of cortisol is vital for homeostasis and adaptive functions, chronic elevation leading to persistent activation can lead to HPA axis dysregulation, which results in hypercortisolism or hypocortisolism.^[7] Hypercortisolism is correlated with mood disorders, cognitive dysfunction, and metabolic abnormalities, while hypocortisolism has been implicated in conditions such as chronic fatigue syndrome and post-traumatic stress disorder.^[8] Besides cortisol, stress impacts other hormonal pathways as well. The autonomic nervous system becomes dysregulated by chronic stress, promoting elevated levels of catecholamines like adrenaline and noradrenaline that are associated with increased cardiovascular risk and immunosuppression.^[9] Moreover, chronic stress also interferes with the hormonal balance of reproductive hormones like estrogen and testosterone, causing menstrual disorders, decreased libido, and infertility.^[10] Likewise, chronic stress-induced dysregulation of insulin and leptin is a cause of metabolic diseases like insulin resistance, obesity, and type 2 diabetes.^[11] Chronic stress was also found to disrupt oxytocin and beta-endorphin levels; hormones that underlie social attachment and pain perception, possibly leading to emotional dysregulation and augmented pain sensitivity.^[12] With these far-reaching consequences, stress management through lifestyle modification, mindfulness, and therapy is essential to ensure hormonal equilibrium and overall health. Non-pharmacological interventions like meditation, yoga, and social support have been found to be effective in re-establishing hormonal homeostasis and preventing stress-induced dysfunction.^[13]

Meditation is an established mind-body technique that has been demonstrated to effectively lower stress, improve mental health, and stimulate the release of neurochemicals related to happiness, like oxytocin and beta-endorphins. Meditation has been shown to decrease HPA axis activity and thereby lower cortisol release and decrease stress reactivity.^[14] Hormones such as endorphins, oxytocin, dopamine, and serotonin – collectively termed “happiness hormones” – play crucial roles in regulating mood, fostering social bonding, and alleviating stress. Mindfulness meditation has been shown to strongly decrease cortisol, creating a condition of relaxed physiological responding.^[15] Other meditation-based interventions, like Loving-Kindness and compassionate approach, have been associated with higher oxytocin levels that are involved in positive emotions and social connection.^[16] Additionally, meditation activates the release of beta-endorphins that are typically involved in improving mood and decreasing stress.^[17]

Heartfulness (HFN) meditation, a modern practice rooted in ancient yogic traditions, offers a unique approach to improving mental health. Unlike conventional meditative techniques, HFN incorporates the concept of yogic transmission – a subtle energy believed to aid in achieving deeper states of relaxation and expanded consciousness. The practice focuses on connecting with the “light within the heart,” fostering a sense of inner calm, clarity, and emotional transformation. This meditative approach has garnered attention for its ability to reduce stress, improve emotional regulation, and enhance personal growth.

In this study, a randomized controlled trial was conducted to evaluate the perception of the meditation experience using HFN meditation techniques. Therefore, the objective of this study was to understand the ability of HFN meditation to modulate the endocrine system through control of the secretion of vital hormones and neurotransmitters associated with happiness and stress management. This research presents scientific findings that indicate that HFN meditation enhances the secretion of oxytocin and beta-endorphins and decreases cortisol levels, hence enhancing emotional strength and mitigating the adverse effects of chronic stress.

2. Methodology

2.1. Study design

A prospective randomized controlled trial was conducted to assess the changes in happy hormones. The participants were randomized into 2 groups-experimental group, where participants were given the intervention of HFN meditation (with yogic transmission) and a control group, where the intervention of meditation was not given (as depicted in Figure 1). Ethical approval (MGM/DCH/IEC/IN/SBS/47/01/2024) was obtained from IEC MGMDCH. This study was also registered in the Clinical Trial Registry of India (CTRI)-CTRI/2024/09/073920 and it was conducted as per the guidelines of Declaration of Helsinki.

Figure 1.

Study design showing random allocation, pre-crossover (phase 1), postcrossover (phase 2), and assessments using MEDEQ, PANAS, and biomarkers. MEDEQ = Meditation Depth Questionnaire.

2.2. Study population

All the students (aged 18 to 25 years) under constituent units of MGMIHS, Navi Mumbai were approached. An email was sent from the Department of Medical Biotechnology, MGMSBS, MGMIHS, Kamothe, Navi Mumbai, to the participants. Interested participants with no previous meditation experience were included in the study. A convenience sample of interested subjects was randomized into 2 groups (experimental and control) prior to the start of study. Participants of experimental group and control group were provided with the electronic informed consent form after completing the initial electronic questionnaire to assess their eligibility. All willing subjects were screened for inclusion and exclusion criteria as follows-

• (i)Inclusion criteria: adults (both male and female) above 18 years of age who were willing to participate in the study.
• (ii)Exclusion criteria: Participants who were unwilling to participate in the study, and participants who were unable to sit for 30 minutes at a stretch due to physical or mental conditions. Subjects with medical conditions in which a blood draw would be contraindicated (e.g., severe anemia), active use of marijuana, opioids, or related drugs, and participants currently living outside of the country were also excluded from this study.

2.3. Intervention

The intervention phase was carried out at MGMSBS MGMIHS, Kamothe, Navi Mumbai. Participants were randomly divided into 2groups Group A (35 participants) experimental group followed a structured meditation program of HFN from a certified trainer, while Group B (35 participants) served as control group, who didn’t receive meditation during the first phase.

• A.Initial session: Group A participants, assigned to a HFN trainer, underwent a HFN introductory session for 3 consecutive days offline. During these sessions, participants closed their eyes and rested their attention on the source of divine light within their hearts for approximately 30 minutes. The HFN trainer meditated with the participants and facilitated yogic transmission.^[18] They continued this process for 1 month (weekly thrice online and once in 15 days through offline sessions). Group B participants, assigned to the control group, did not undergo any HFN session. Both group participants filled out the Meditation Depth Questionnaire (MEDEQ) and Positive and Negative Affect Schedule (PANAS) questionnaires at the beginning. After 1 month, biomarker analysis and MEDEQ/PANAS questionnaire analysis were conducted.
• B.Repeat session with crossover: After the initial session, Group A participants were not given HFN meditation with a certified HFN trainer. However, they were instructed to continue the practice through the HeartsApp. And during this phase Group B participants were assigned to a certified HFN trainer. These participants followed the HFN protocol with the help of a certified trainer.^[18] During this time, the HFN trainer meditated with the participants and facilitated yogic transmission. After completing the session, both group participants underwent postbiomarker analysis and MEDEQ/PANAS questionnaire analysis.

2.4. Assessment protocol

1. Baseline assessment: All participants completed pre-intervention testing, which included self-reported measures using the MEDEQ and PANAS questionnaires.

2. Post-intervention assessment: Following 1 month of intervention, all participants underwent post-intervention testing which included biochemical assay (serum oxytocin, serum β-endorphin and serum cortisol levels) and self-reported measures of Meditation Depth Index (MEDI) and PANAS questionnaires.

3. Final assessment: At the end of the crossover phase, both groups underwent post-crossover biochemical assessments and repeated administration of the MEDI and PANAS questionnaires. Data from baseline, pre-crossover (i.e., Day 30), and post-crossover (i.e., Day 60) assessments were analyzed to assess the impact of the HFN meditation intervention.

2.5. Assessment tools

• 1]MEDEQ Questionnaire: The MEDEQ is a self-report tool developed by Prion et al (2001) to assess the subjective depth of a person’s meditative experience during a specific session. It is widely used in meditation and mindfulness research and provides a standardized way to quantify meditative depth, making it useful for evaluating the effectiveness of different meditation practices or interventions.^[19]
• 2]PANAS Questionnaire: The PANAS is a widely used self-report questionnaire developed by Watson, Clark, and Tellegen (1988) to measure 2 primary dimensions of mood: positive affect (PA) and negative affect (NA). It consists of 20 items – 10 assessing positive emotions (e.g., enthusiastic, alert) and 10 assessing negative emotions (e.g., distressed, upset) – which participants rate based on how they feel over a specified time frame (e.g., “right now,” “past week”). Each item is rated on a Likert scale, typically from 1 (very slightly or not at all) to 5 (extremely).^[20]
• 3]Biomarkers: Fasting blood samples (3 mL) were collected from all participants between 8:00 and 8:30 am under aseptic conditions at 2 time points – before and after crossover (at day 30 and day 60). The samples were drawn into plain vacutainer tubes and transported to the MGM Central Research Laboratory, Department of Medical Biotechnology, Kamothe, Navi Mumbai, for analysis. Samples from both experimental and control group participants were collected Blood samples were centrifuged at 3000 rpm for 10 minutes, and serum was separated within 30 minutes of collection, then stored at −80°C until further analysis. Biochemical markers such as serum oxytocin, endorphin and cortisol were assessed using ELISA-based assays. The following kits were used in accordance with the manufacturers’ protocols: Human Oxytocin ELISA Kit (Elabscience, Houston; sensitivity: 9.38 pg/mL, detection range: 15.63–1000 pg/mL), Human Beta-endorphin ELISA Kit (Elabscience; sensitivity: 9.38 pg/mL, detection range: 15.63–1000 pg/mL) and Human Cortisol ELISA Kit (Elabscience; sensitivity: 2.92 ng/mL, detection range: 6.25–400 ng/mL). Each participant visited twice for data collection and blood sampling.

2.6. Sample size calculation

Stratified random sampling method was used for the selection of the sample. The representative sample was selected from interested volunteers. G*Power software was used to determine the sample size by applying a t-test family to compare the difference between 2 independent means (two groups: experimental and control group). An expected effect size of d = .75 (P = .05 and statistical power = .80) was taken, based on the results of a brief mindfulness session on several biological markers, with a 20% dropout^[21] and Sample size group 1 = 29 Sample size group 2 = 29 was obtained. After assuming a 20% dropout rate, 35 participants were enrolled in the experimental group and 35 in the control group; total sample size = 70.

2.7. Randomization

Computer generated randomization was done by a blinded research assistant and participants were allocated into experimental group and control group. The participants were blinded from the study.

2.8. Statistical analysis

Statistical analysis was carried out using the software IBM SPPS software version 26.0. The distribution of data was determined using Kolmogorov–Smirnov statistics (P > .05 indicates normality is assumed) and continuous variables were represented as mean (SD). Repeated measures analysis of variance (RM-ANOVA) statistics was used to compare the mean outcomes between the experimental group and control group from baseline till post-crossover phase. The between-group outcomes were analyzed using Mann–Whitney U test and Wilcoxon Signed-Rank Test for within group comparisons. The correlation between variables was analyzed using the Spearman test.

3. Results

The present study aimed to evaluate the effects of a 2-month HFN meditation intervention on happy hormones (oxytocin and β-endorphin) and cortisol (stress biomarker). The participants didn’t report any harm during the intervention. The Consolidated Standards of Reporting Trials (CONSORT) flowchart of participants enrolled in this study is depicted in Figure 2.

Figure 2.

Consort flow chart of the study.

3.1. Demographic profile

The sociodemographic characteristics of 60 participants aged 18 to 25 years (M = 22.29, SD = 2.98), with 48 (80%) females and 12 (20%) males are presented in Table 1. Regarding residence, 20 (33.33%) lived in their own house, 19 (31.66%) in rented homes, 9 (15%) as paying guests, and 12 (20%) in hostels. The mean BMI was 23.77 (SD = 5.21), and participants reported an average of 6.45 hours of sleep daily (SD = 1.07). Diet patterns showed 24 (40%) were vegetarian, 36 (60%) non-vegetarians, with no vegans. Junk food consumption varied: 12 (20%) daily, 20 (33.33%) twice a week, 12 (20%) thrice a week, 10 (16.66%) once a week, 4 (6.66%) every 15 days, and 2 (3.33%) monthly. Lifestyle habits included 28 (46.66%) exercising, and 21 (35%) practicing yoga. Participants spent an average of 6.45 hours daily on the internet (SD = 1.07), using mobile devices (36, 60%) and laptops (24, 40%).

Table 1.

Sociodemographic characteristics of all participants.

Characteristics	No. of Participants (n)	Percentage (%)
Age
Mean (SD)	22.29 (2.98)	–
Range	18–25
Gender
Male	12	20
Female	48	80
Religion
Hindu	48	80
Islam	9	15
Buddhism	2	3.33
Christianity	1	1.66
Place of residence
Own house	20	33.33
Rented home	19	31.66
Paying guest	9	15
Hostel	12	20
BMI
Mean (SD)	23.77 (5.21)	–
How many hours do you sleep in a day?
Mean (SD)	6.45 (1.07)	–
Diet pattern
Vegetarian food	24	40
Vegan food	0	0
Non-vegetarian food	36	60
Frequency of junk food in a week
Every day	12	20
Twice a week	20	33.33
Thrice a week	12	20
Once a week	10	16.66
Every 15 d	4	6.66
Once a month	2	3.33
Do you exercise?
Yes	28	46.66
No	32	53.33
Do you perform Yoga?
Yes	21	35
No	39	65
How many hours do you spend on Internet per day?
Mean (SD)	6.45 (1.07)	-
The device used for Internet
Mobile	36	60
Laptop	24	40
What do you do most on your device?
Reading on mobile	16	26.66
Watching movies on mobile	7	11.66
Playing games on mobile	6	10
Using social media on mobile	7	11.66
Reading on laptop	6	10
Watching movies on laptop	7	11.66
Playing games on laptop	9	15
Using social media on laptop	2	3.33
Any kind of addiction
Alcohol consumption	–
Smoking	–
Tobacco consumption	–
None	60	100
Any gap between study?
Yes	11	18.33
No	49	81.66
Residence
With family	26	43.33
With friends	14	23.33
Without family	10	16.66
Nuclear	10	16.66

3.2. Qualitative assessment

3.2.1. MEDEQ analysis

The meditation depth of participants of both groups (Experimental & Control) was evaluated at 3 time points – Days 1, 3, and 30, as shown in Table 2.

Table 2.

MEDEQ scores between experimental group and control group at different time intervals before crossover.

MEDEQ	Day = 1	Day = 3	Day = 30
Experimental group	57 (SD = 2.325)	64.407 (SD = 1.605)	83.481 (SD = 1.366)
Control group	56.703 (SD = 1.626)	58.407 (SD = 1.820)	59.963 (SD = 2.846)

MEDEQ = Meditation Depth Questionnaire.

The RM-ANOVA analysis was carried out to determine if there was a significant difference between the mean MEDEQ scores between these 2 groups across 3 time points. These results are shown in Table 3. There was a significant main effect of intervention on meditation depth (F = 13.478, P = .001, r = 0.333) suggesting that the intervention leads to meaningful changes. Additionally, time also has a highly significant effect (F = 11.650, P = .000, r = 0.301), implying that the MEDEQ scores changes significantly over different time points. The interaction between intervention and time was also found to be significant (F = 7.813, P = .001, r = 0.224), indicating that the effect of the intervention varies across time points.

Table 3.

RM-ANOVA analysis of MEDEQ scores between experimental group & control group before crossover.

Source	Type III sum of squares	Df	Mean square	F	Sig.	Effect size (r)
Intervention	4148.308	1	4148.308	13.478	.001	0.333
Time	6501.160	2	3250.580	11.650	.000	0.301
Intervention × time	4100.611	2	2050.305	7.813	.001	0.224

MEDEQ = Meditation Depth Questionnaire, RM-ANOVA = repeated measures analysis of variance.

Pairwise comparisons were also carried out to determine which time points showed the difference (as shown in Table 4). The comparisons between Days 1 and 30 (M = –14.87, 95% CI [–23.61,–6.13], P = .001) and between Days 3 and 30 (M=–10.32, 95% CI [–17.49,–2.69], P = .006) yield statistically significant differences. These findings suggest a significant change in the MEDEQ scores over a longer duration, with differences emerging between the short-term (Days 1 and 3) and the long-term (Day 30) intervals.

Table 4.

Pairwise comparisons of MEDEQ scores between experimental group and control group before crossover.

Time intervals	Mean difference (M)	Sig.†	95% CI for difference†
			Lower bound	Upper bound
Day = 1
Day = 3	−4.556	.437	−12.316	3.204
Day 30	−14.870*	.001	−23.609	−6.131
Day = 3
Day = 1	4.556	.437	−3.204	12.316
Day 30	−10.315*	.006	−17.943	−2.687
Day = 30
Day = 1	14.870*	.001	6.131	23.609
Day = 3	10.315*	.006	2.687	17.943

MEDEQ = Meditation Depth Questionnaire.

^†Adjustment for multiple comparisons: Bonferroni.

^*The mean difference is significant at the .05 level.

After 30 Days of intervention in the experimental group, the control group participants were given the intervention of HFN meditation for 1-month (crossover). The mean scores after crossover are mentioned in Table 5. The MEDEQ scores of control group significantly increased from 59.70 (SD = 1.71) to 72.41 (SD = 3.27) (P = .006) after crossover. However, the MEDEQ scores in the experimental group did not differ significantly during this period (P = .580). This suggests that the HFN meditation intervention was effective in improving MEDEQ scores in those who had not previously received it (control group), indicating a positive impact of the intervention.

Table 5.

MEDEQ scores before and after crossover in experimental and control group.

MEDEQ	Experimental group	P-value	Control group	P-value
Before crossover	83.48 (SD = 1.54)	.580	59.963 (SD = 2.846)	.006
After crossover	84.67 (SD = 2.83)		72.41 (SD = 1.27)

MEDEQ = Meditation Depth Questionnaire.

3.2.2. PANAS positive analysis

The positive effect of intervention on mood of participants of both groups (Experimental & Control) was evaluated at 3 time points – Days 1, 3, and 30, as shown in Table 6.

Table 6.

PANAS (positive) scores between experimental group and control group before crossover.

PANAS-POS	Day = 1	Day = 3	Day = 30
Experimental group	30.00 (SD = 3.431)	30.48 (SD = 3.333)	38.48 (SD = 3.079)
Control group	31.89 (SD = 3.363)	31.96 (SD = 3.549)	31.78 (SD = 3.500)

PANAS = Positive and Negative Affect Schedule.

The RM-ANOVA analysis was carried out to determine if there was a significant difference between the mean PANAS positive scores between these 2 groups across 3 time points. These results are shown in Table 7. The main effect of intervention on PA on mood (F = 0.81, P = .377, r = 0.03) was not statistically significant. But, time showed a highly significant effect on positive scores (F = 7.06, P = .002, r = 0.21), implying that these scores change significantly over different time points. The interaction between intervention and time was also found to be highly significant (F = 10.80, P ≤ .000, r = 0.29), indicating that the effect of the intervention varies across time points.

Table 7.

RM-ANOVA analysis of PANAS (positive) scores between experimental group and control group before crossover.

Source	Type III sum of squares	Df	Mean square	F	Sig.	Effect size
Intervention	50.0	1	50.0	0.81	.377	0.03
Time	591.49	2	295.75	7.06	.002	0.21
Treatment × time	634.481	2	317.241	10.80	.000	0.29

PANAS = Positive and Negative Affect Schedule, RM-ANOVA = repeated measures analysis of variance.

Pairwise comparisons were also conducted to identify which time points showed the significant differences (as shown in Table 8). It was observed that the time intervals Days 1 and 30 (M = –8.481, 95% CI [–12.03,–4.93], P ≤ .000) and Days 3 and 30 (M = –8.000, 95% CI [–13.52,–2.48], P = .003) showed statistically significant differences. These findings suggest a significant change in the PANAS positive scores over time, with the most pronounced effect observed by Day 30.

Table 8.

Pairwise comparisons of PANAS-(positive) scores between experimental group and control group before crossover.

Time intervals	Mean difference (M)	Sig.†	95% CI for difference†
			Lower bound	Upper bound
Day = 1
Day = 3	−0.481	.000	−4.073	3.110
Day = 30	−8.481*	.000*	−12.028	−4.935
Day = 3
Day = 1	.481	1.000	−3.110	4.073
Day = 30	−8.000*	.003*	−13.523	−2.477
Day = 30
Day = 1	8.481*	.000*	4.935	12.028
Day = 3	8.000*	.003*	2.477	13.523

PANAS = Positive and Negative Affect Schedule.

^†Adjustment for multiple comparisons: Bonferroni.

^*The mean difference is significant at the .05 level.

The mean scores after crossover are mentioned in Table 9. The PANAS positive scores of control group significantly increased from 31.78 (SD = 3.50) to 36.81 (SD = 3.69) (P = .004) after crossover. However, these scores in the experimental group did not differ significantly during this period (P = .409). This suggests that the HFN meditation intervention was effective in improving PANAS scores in those who had not previously received it (control group), indicating a positive impact of the intervention.

Table 9.

PANAS (positive) scores between before and after crossover.

PANAS positive	Experimental group	P-value	Control group	P-value
Before crossover	37.14 (SD = 3.92)	.409	31.78 (SD = 3.50)	.004
After crossover	39.44 (SD = 3.01)		36.81 (SD = 3.69)

PANAS = Positive and Negative Affect Schedule.

3.2.3. PANAS negative analysis

The NA of intervention on mood of participants of both groups (experimental and control) was evaluated at 3 time points – Days 1, 3, and 30, as shown in Table 10.

Table 10.

PANAS (negative) scores between experimental group & control group before crossover.

PANAS-NEG	Day = 1	Day = 3	Day = 30
Experimental group	26.63 (SD = 3.239)	23.85 (SD = 3.696)	18.93 (SD = 3.220)
Control group	26.44 (SD = .653)	25.22 (SD = 3.706)	25.67 (SD = 3.102)

PANAS = Positive and Negative Affect Schedule.

The RM-ANOVA analysis was carried out to determine if there was a significant difference between the mean PANAS negative scores between these 2 groups across 3 time points. These results are shown in Table 11. It was observed that the effect of intervention on NA on mood was statistically significant (F = 6.72, P = .015, r = 0.21), suggesting that the intervention leads to meaningful changes. Additionally, time has a highly significant effect (F = 5.67, P = .006, r = 0.18), implying that these scores changes significantly over different time points. The interaction between intervention and time was also found to be significant (F = 3.13, P = .052, r = 0.11), indicating that the effect of the intervention varies across time points.

Table 11.

RM-ANOVA analysis of PANAS (negative) scores between experimental group and control group before crossover.

Source	Type III sum of squares	Df	Mean square	F	Sig.	Effect size
Treatment	282.691	1	282.70	6.72	.015	0.21
Time	486.086	2	243.04	5.67	.006	0.18
Treatment × time	356.531	2	178.26	3.13	.052	0.11

PANAS = Positive and Negative Affect Schedule, RM-ANOVA = repeated measures analysis of variance.

Pairwise comparisons were also carried out to determine which time points showed the difference (as shown in Table 12). It was observed that the time intervals Days 1 and 30 (M = 7.704, 95% CI [–3.81, 11.60], P = .000) and Days 3 and 30 (M = 4.296, 95% CI [0.74, 9.11], P = .017) showed statistically significant differences. These findings suggest a significant change in the PANAS negative scores over a longer duration, with differences emerging between the short-term (Days 1 and 3) and the long-term (Day 30) intervals.

Table 12.

Pairwise comparisons of PANAS-(negative) scores between experimental group and control group before crossover.

Time intervals	Mean difference (M)	Sig.†	95% CI for difference†
			Lower bound	Upper bound
Day = 1
Day = 3	2.778	.481	−2.142	7.697
Day = 30	7.704*	.000*	3.805	11.603
Day = 3
Day = 1	−2.778	.481	−7.697	2.142
Day = 30	4.926*	.017*	0.741	9.111
Day = 30
Day = 1	−7.704*	.000*	−11.603	−3.805
Day = 3	−4.926*	.017*	−9.111	−0.741

PANAS = Positive and Negative Affect Schedule.

^†Adjustment for multiple comparisons: Bonferroni.

^*The mean difference is significant at the .05 level.

The mean scores after crossover are mentioned in Table 13. The PANAS negative scores of control group significantly decreased from 25.67 (SD = 3.10) to 19.00 (SD = 3.64) (P ≤ .000) after crossover. However, these scores in the experimental group did not differ significantly during this period (P = .508). This suggests that the HFN meditation intervention was effective in improving PANAS scores in those who had not previously received it (control group), indicating a positive impact of the intervention.

Table 13.

PANAS (negative) scores between before and after crossover.

PANAS negative	Experimental group	P-value	Control group	P-value
Before crossover	18.29 (SD = 3.99)	.508	25.67 (SD = 3.10)	.000
After crossover	18.85 (SD = 3.26)		19.00 (SD = 3.64)

PANAS = Positive and Negative Affect Schedule.

3.3. Quantitative assessment

3.3.1. Happy hormone – oxytocin

The biological hormones were also assessed to study the impact of the intervention of HFN meditation. It was observed that the level of happy hormone oxytocin was higher in the experimental group (838.18, SD = 12.477) as compared to the control group (750.00, SD = 16.59, P = .003, r = 0.355) at day 30. After the crossover, the oxytocin levels in both the experimental (897.11, SD = 11.56) and control group (825.88, SD = 10.44, P = .003, r = 0.355) increased significantly at day 60 (as shown in Figure 3). Additionally, there was a significant increase in the oxytocin levels of control group after crossover (825.88, SD = 10.44) as compared to before crossover (750.00, SD = 16.59, P = .001, r = 0.588).

Figure 3.

Comparison of oxytocin levels between experimental group and control participants before and after crossover. Significant increases (P < .01) in oxytocin levels were observed in the experimental group compared to the control group at both time points. ** P ≤ .01.

3.3.2. Happy hormone – endorphin

The level of happy hormone endorphin was significantly higher in the experimental group (692.83, SD = 17.31) as compared to the control group (598.95, SD = 12.29, P = .003, r = 0.357) at day 30. After the crossover, the endorphin levels in both the experimental (748.59, SD = 13.95) and control group (693.78, SD = 12.36. P = .05, r = 0.235) increased significantly at day 60 (as shown in Figure 4). Additionally, there was a significant increase in the endorphin levels of control group after crossover (693.78, SD = 12.36) as compared to before crossover (598.95, SD = 12.29, P = .002, r = 0.563).

Figure 4.

Comparison of endorphin levels between experimental group and control participants before and after crossover. Significant increases in endorphin levels were observed in the experimental group compared to the control group at both time points. * P ≤ .05, ** P ≤ .01.

3.3.3. Stress hormone – cortisol

The biological stress hormone cortisol was also assessed to study the impact of the intervention of HFN meditation. It was observed that the level of cortisol decreased significantly in the experimental group (158.81, SD = 15.93) as compared to the control group (292.36, SD = 16.81, P = .000, r = 0.661) at day = 30 (as shown in Figure 5). Additionally, the cortisol levels decreased significantly in the control group after crossover (200.26, SD = 15.61, P = .000, r = 0.658).

Figure 5.

Comparison of cortisol levels between experimental group and control participants before and after crossover. Significant reduction in cortisol levels was observed in the experimental group compared to the control group at both time points.*** P ≤ .001.

After the crossover phase, both happy hormones and stress biomarkers-oxytocin, endorphin and cortisol were found to be significantly altered after the intervention of HFN meditation was given to the control group. The levels of happy hormones significantly increased, whereas the level of cortisol, significantly decreased as depicted in Figure 6.

Figure 6.

Comparison of all serum biomarkers in control participants before and after crossover. Significant increases in oxytocin and endorphin levels and significant reduction in cortisol levels were observed in the control group between both time points. *** P ≤ .001.

3.4. Correlation analysis

The correlation between the happy hormones and stress hormone was determined. A significant negative correlation was observed between oxytocin and endorphin with cortisol respectively, as shown in Table 14.

Table 14.

Correlation between happy hormones and cortisol.

Happy hormones	Cortisol (r)	P-value
Endorphin	−0.076	.584
Oxytocin	−.356**	.008

The correlation between the happy hormones and psychometric measures was also determined, as shown in Tables 15 and 16. A significant positive correlation was observed between endorphin and MEDI as well as PANAS positive; whereas endorphin and oxytocin both showed negative correlation with PANAS negative. Oxytocin also showed significant positive correlation with MEDI.

Table 15.

Correlation between happy hormones and MEDI.

Happy hormones	MEDI (r)	P -value
Endorphin	0.336*	.013
Oxytocin	0.306*	.025

MEDI = Meditation Depth Index.

Table 16.

Correlation between happy hormones & PANAS.

Happy hormones	PANAS positive (r)	P-value	PANAS negative (r)	P-value
Endorphin	.459**	.000	−0.079	.568
Oxytocin	0.262	.056	0.013	.926

PANAS = Positive and Negative Affect Schedule.

4. Discussion

This study was conducted to understand the changes in happy hormones such as oxytocin and β-endorphins, stress hormone – cortisol, as well as psychometric aspects such as meditation depth (MEDEQ) and positive and negative emotions (PANAS), after the intervention of HFN meditation; comparing outcomes between the experimental and control groups. The key findings of this study are novice meditators who practiced HFN showed higher MEDI and PANAS scores compared to the control group. The experimental group had significantly elevated oxytocin and β-endorphin levels compared to the control group. Serum cortisol levels, a stress biomarker, also decreased in the experimental group compared to the control group. After crossover, the control group participants also showed improvements in these parameters.

At the start of the intervention period, participants in the experimental group reported a moderate level of meditation experience. Following the intervention of HFN meditation, there was a significant increase in their scores, indicating a significant deepening of the meditative experience. The control group also demonstrated a statistically significant improvement in their meditation depth experience, only after they received the intervention after the crossover phase. Expanding on this, Hölzel et al.,2007 examined the relationships between meditation depth, absorption, meditation practice, and mindfulness using the MEDEQ questionnaire. It was observed that meditators who had more practice of meditation had deeper meditations as compared to those who had lesser experience in meditation. Their analysis of 251 meditators also found that absorption had a stronger influence on meditation depth (path coefficient: 0.48) than meditation practice (0.21).^[22] Similarly, a study comparing the effects of HFN meditation in long-term meditators, short-term meditators, and control group (non-meditators), showed increased MEDEQ scores among long-term meditators and short-term meditators as compared to the non-meditators.^[23] Deep meditative state leads to suppression of global vagal modulation and enhancement of sympathetic and baroreflex activity, which also influences the breathing rhythm.^[24] This leads to improved autonomic regulation, leading to a relaxed and balanced state; which further leads to decrease in stress levels and improvement in happy hormone levels.

The positive and NA of the intervention was assessed using the PANAS tool. These results indicated that before the intervention, participants in the experimental group reported moderate levels of both positive and NA. Following the intervention, there was a clear increase in PA and a significant reduction in NA. In the control group, baseline affect scores remained relatively unchanged until they received the intervention, after which improvements were observed in both positive and NA measures. Likewise, in a study by de Bruin et al (2020) examined the impact of a 6-week program combining exercise, yoga, and mindfulness on work-related stress. The PANAS questionnaire was used to assess changes in emotional states. The findings revealed significant emotional improvements, with PA increasing from 27.04 (SD = 5.21) to 31.71 (SD = 5.08) and NA decreasing from 26.17 (SD = 6.45) to 21.67 (SD = 6.57) post-intervention.^[25] Similarly, another clinical trial by La Torre et al (2020) evaluated the effects of a combined yoga and mindfulness program on healthcare workers, which led to a significant decrease in NA scores; from a median of 16 pre-intervention to 10 post-intervention (P < .001), suggesting that integrating yoga and mindfulness can effectively reduce negative emotional states in high-stress professions.^[26] These findings suggest that HFN meditation plays a significant role in enhancing emotional well-being, as reflected in increased meditation depth and PA, along with a decrease in negative emotions.

Our study also demonstrated that HFN meditation had a positive impact on key biomarkers, including serum oxytocin, β-endorphin and cortisol levels, which are associated with emotional well-being and stress regulation. Our study examined the levels of serum β-endorphin, which is a hormone associated with stress regulation, mood enhancement, and overall well-being. It was observed that the level of happy hormone endorphin was significantly higher in the experimental group as compared to the control group at day 30. After the crossover, the endorphin levels in both the experimental and control group increased significantly at day 60. There was a significant increase in the endorphin levels of control group after crossover as compared to before crossover. This implies that the intervention of HFN meditation was able to increase the production of endorphin in these participants. The same was observed by Gagrani et al, 2018. They carried out an intervention of meditation for 6 weeks, along with the standard care (intervention group) and only standard care treatment in the control group. They selected patients suffering from glaucoma as this neurodegenerative disorder is found to have an association with quality of life as well as stress. They found that the mean endorphin level was higher in intervention group (33, SD = 5.52pg/mL to 43.27pg/mL, P < .0001), when compared with levels in the control group (34.78, SD = 4.1pg/mL to 36.33, SD = 4.07pg/mL, P = .27). This further implies that meditation can be used as an adjunct therapy in glaucoma patients.^[27] Similarly, a study by d’Arma et al (2022) examining the impact of non-pharmacological interventions on β-endorphin levels in patients with multiple sclerosis found a significant increase over the course of the intervention. Baseline β-endorphin levels were recorded at 113.4 pg/mL, rising to 142.0 pg/mL by the end of the study, with a statistically significant difference (P < .02). These findings suggest that non-pharmacological approaches may play a beneficial role in enhancing β-endorphin production in individuals with multiple sclerosis.^[28] Another study by Bidari et al (2016) aimed to investigate the effects of strenuous exercise on β-endorphin (β-END) levels in fibromyalgia patients compared to healthy individuals. The results showed a significant increase in β-END levels following exercise in both groups. In the fibromyalgia group, pre-exercise β-END levels were 90.12 (SD = 20.91) µg/mL, which increased to 179.80 (SD = 28.57) µg/mL postexercise (P < .001) and in the healthy group, pre-exercise β-END levels were 122.07 (SD = 28.56) µg/mL, rising to 246.55 (SD = 29.57) µg/mL postexercise (P < .001).^[29] However, a study in 1999 by Øktedalen et al showed dissimilar results. They found that even though intense physical training significantly increased beta-endorphin levels and reduced pain perception, Association for Comprehensive Education in Meditation did not have the same effect.^[30] Although another research studying these 2 kinds of activities (running and meditation) showed significant mood elevation after intervention, moreover the difference between the groups was not statistically significant, indicating that even though these activities are metabolically different, their effect was similar on these hormones.^[31] Beta-endorphins are known to be natural painkillers. They are produced in the pituitary gland and central nervous system, and can bind to opioid receptors in the brain, thereby reducing the perception of pain and promoting well-being.^[32] During HFN meditation, we found an increase in endorphin levels, indicating a sense of calmness and stress reduction. As this hormone has morphine like activity, its release may have mood elevating activity.^[33]

Oxytocin was also examined in our study. Oxytocin is often associated with loneliness; lower levels are associated with higher feelings of loneliness.^[34] In our study, it was observed that the level of oxytocin was higher in the experimental group as compared to the control group at day 30. After the crossover, the oxytocin levels in both the experimental and control group increased significantly at day 60. Additionally, there was a significant increase in the oxytocin levels of control group after crossover as compared to before crossover. This implies that the intervention of HFN meditation was able to increase the production of oxytocin in these participants. Similarly, in a study conducted on Human Immunodeficiency Virus patients, oxytocin was found to be double in participants who were spiritually transformed as compared to those who were not.^[35] In another study conducted on healthy population of a religious community, they found high spirituality levels (M = 5.38, SD = .55) and oxytocin levels to be 6.07 pg/mL (SD = 2.56).^[36] A previous study on mindfulness meditation by Aygün et al (2024) demonstrated a significant increase in serum oxytocin levels in the meditation group compared to the control group, rising from 8.81 ng/mL to 18.21 ng/mL (P < .038). This suggests that meditation leads to a greater elevation in oxytocin levels in the intervention group.^[37] Oxytocin is associated with feelings of stress reduction, social bonding,^[38] trust,^[39] attachment^[40] and emotional empathy between human beings.^[41] Research indicates that oxytocin plays a crucial role in reducing pain sensitivity by enhancing mood. Meditation practices contribute to autonomic nervous system regulation by enhancing parasympathetic activity and reducing sympathetic responses. As a result, consistent practice of HFN meditation fosters relaxation, promotes emotional balance, and plays a vital role in stress reduction.^[37]

The biological stress hormone cortisol was also assessed to study the impact of the intervention of HFN meditation. It was observed that the level of cortisol decreased significantly in the experimental group as compared to the control group at day 30. Additionally, the cortisol levels also decreased significantly in the control group after crossover. A study was conducted on glaucoma patients to study the effect of meditation intervention. Glaucoma is a neurodegenerative disease which is closely associated with stress and adverse quality of life. The intervention group participants underwent 45 minutes of meditation daily for 6 weeks in addition to standard medical treatment while controls received only standard medical treatment. Mean serum cortisol decreased significantly in intervention group (497, SD = 46.37ng/mL to 447, SD = 53.78ng/mL, P = .01) as compared to control group (519.75, SD = 24.5 to 522.58, SD = 26.63ng/mL, P = .64), indicating that meditation leads to reduction in the stress hormone cortisol.^[27] Another interesting study evaluated the effects of web-based HFN meditation on dehydroepiandrosterone sulfate (DHEA-S); a hormone secreted by adrenal cortex that shows anticortisol effects. The levels of this hormone were significantly elevated in the HFN meditation group as compared to the control group. The practice of HFN may promote a state of deep mental relaxation, which could contribute to elevated levels of DHEA-S.^[42] Similarly, a study on COVID-19 patients (aged 18–50 years) showed a significant decrease in blood parameters like circulating cortisol, oxidative stress biomarkers and inflammatory biomarkers in participants of the HFN meditation group as compared to the control group (who received relaxation intervention), after an app-based intervention of 4 weeks. This indicates that HFN meditation can be used as a non-pharmacological supportive approach to accelerate the recovery process in patients who have completed the COVID-19 treatment regimen.^[43] These findings altogether reinforce the growing body of evidence that meditation serves as a valuable non-pharmacological approach to stress reduction and overall well-being.

We also examined the correlation of these hormones with MEDEQ and PANAS. It is observed that oxytocin is associated with both loneliness and higher levels of cortisol.^[44,45] In our study also, we found a significant negative correlation between oxytocin and cortisol levels; suggesting that oxytocin helps reduce cortisol levels by promoting a sense of calmness. Similarly, a pilot study by Li Yet al.,2019 examined the relationship between oxytocin and cortisol in 3 groups: individuals with post-traumatic stress disorder (PTSD) only, those with both PTSD and depression (PTSD-D), and trauma-exposed resilient controls. The study found an inverse relationship between oxytocin and cortisol levels across all groups.^[46] We also observed a negative correlation between β-endorphin and cortisol levels, indicating a potential role of β-endorphins in mood enhancement and calming effects. A previous study on HFN meditation showed that meditation depth index was positively correlated with mindfulness and negatively with anxiety and impulsiveness.^[23] Similar to this study, our study also showed a significant positive correlation of meditation depth index and PANAS positive with the happy hormones oxytocin and endorphin; suggesting that deeper meditative experiences are associated with increases in these happy hormones reflecting greater emotional positivity and relaxation. However, the positive correlation between PANAS positive and oxytocin was not statistically significant. A negative correlation was observed between negative trait of PANAS and endorphin; suggesting that higher endorphin levels may help reduce negative emotional states. Whereas, a positive correlation was found between PANAS negative and oxytocin, although these were not significant statistically. Overall, these results reinforce the psychophysiological benefits of contemplative practices such as HFN meditation and highlight the relevance of hormonal markers in assessing emotional states.

Therefore, meditation is a practice that has been identified as a potential preventive tool for applications in the field of mental health disorders, as it has shown promising effects in reducing measures like anxiety, stress and depression.^[47–51] Accordingly, this study also contributes to the growing body of evidence supporting that meditation (specifically HFN meditation) can be used a preventive tool for managing stress and mood related problems by reducing stress biomarkers and enhancing the levels of happy hormones.

5. Limitations and future directions

While the current study offers compelling evidence on the positive psychophysiological effects of HFN meditation, there are several limitations in this study. Firstly, the sample size was modest, which may limit the generalizability of the findings across diverse populations. Secondly, the short intervention duration (30 days per phase) may not be sufficient to assess the long-term sustainability of observed hormonal and psychological changes. Thirdly, the hormonal biomarkers were only measured at 2 time points (pre- and postintervention), which may not indicate the fluctuations or peak effects over time.

Therefore, future research should address these limitations by incorporating larger and more diverse samples to enhance applicability, employing longer follow-up periods to examine the persistence of hormonal and emotional benefits. It would also be beneficial to explore additional biomarkers such as serotonin, dopamine, or inflammatory cytokines to build a more comprehensive picture of the neurobiological pathways influenced by meditation. Longitudinal studies evaluating the effect of sustained HFN practice on mental health outcomes in clinical populations (e.g., individuals with depression, or chronic disorders) could further validate its utility as a non-pharmacological intervention.

6. Conclusion

Our study highlights the positive impact of HFN meditation on mood-enhancing hormones and psychological stress, promoting relaxation and overall well-being. Our findings indicate significant improvements in biomarkers and psychological health, suggesting its effectiveness as a holistic approach to stress reduction. Regular practice enhances mental clarity, and emotional resilience, supporting a healthier lifestyle and improved mental well-being. Our study highlights that a structured HFN meditation program effectively reduces stress while enhancing relaxation and overall well-being. This is evidenced by increased oxytocin and β-endorphin levels, deeper meditation experiences, and elevated positive emotions, as measured by the MEDEQ and PANAS questionnaires in individuals with no prior meditation experience. These findings reinforce the effectiveness of HFN meditation as a non-pharmacological approach to boosting mood-enhancing hormones. Therefore, our results suggest that the practice of HFN meditation can be used as a non-pharmacological tool to enhance mood and reduce stress levels, as indicated by the elevation of the happy hormones – oxytocin and endorphin – and reduction of the stress hormone cortisol, after the intervention of HFN meditation. Therefore, this practice can be used a potential tool for stress management and mood enhancement.

Acknowledgments

The authors gratefully acknowledge the support and encouragement of Hon’ble Vice Chancellor and Pro-Vice Chancellor of MGMIHS, which enabled the successful execution of this study. Sincere appreciation is extended to all participants for their time and involvement. The authors also express deep gratitude to Dr Kamlesh Patel, Global Guide of Heartfulness, for his inspirational guidance and support.

Author contributions

Conceptualization: Sanjana T. Philip, Jayaram Thimmapuram, Kapil Thakur, Kishore Sabbu, Mansee Thakur.

Data curation: Sanjana T. Philip, Navami Dayal.

Formal analysis: Sanjana T. Philip.

Investigation: Sanjana T. Philip, Samruddhi Surve.

Methodology: Sanjana T. Philip, Poonam Patil.

Project administration: Sanjana T. Philip, Jayaram Thimmapuram, Kapil Thakur, Mansee Thakur.

Supervision: Jayaram Thimmapuram, Kapil Thakur, Kishore Sabbu, Mansee Thakur.

Validation: Sanjana T. Philip, Kapil Thakur, Yogesh Patil, Mansee Thakur.

Visualization: Sanjana T. Philip, Kapil Thakur, Navami Dayal, Yogesh Patil, Mansee Thakur.

Writing – original draft: Sanjana T. Philip, Mansee Thakur.

Writing – review & editing: Kapil Thakur, Kishore Sabbu.