Effect of Rosmarinus Officinalis Essential Oil On Anxiety, Depression, And Sleep Quality

of Rosmarinus


INTRODUCTION
Emotional disorders cause a considerable burden on global health, emerging evidence suggests that levels of stress and anxiety-related symptoms increased during the COVID-19 pandemic. 1,2n fact, previous experiences have shown that infectious disease outbreaks are related to a greater number of mental health problems in survivors and affected communities. 3In this sense, the postpandemic era will probably be characterized by an increase in mental health problems. 4Data suggest that high levels of anxiety, depression symptoms, and sleep problems have been prevalent during pandemic, 5 especially among young adults, showing a consistent increasing trend. 6Literature also indicates that stress may trigger anxiety as well as sleep difficulties and depression. 7In this context, insomnia is one of the most prevalent sleep disorders, defined as a chronic statement of difficulty falling or staying asleep. 8In this regard, insomnia, anxiety, and depression share common etiological processes, however, evidence indicates a cyclical relationship, that is, sleep disorders impulse the development of depression 9 as well as sleep disorders induce depression. 10Therefore, as an investigation may sustain, improving sleep quality could prevent, manage, and alleviate stress and depression symptoms. 11wadays, there is a growing trend to use herbal medicines as an alternative to treat different ailments.This is in accordance with the belief that all natural products are safer and cheaper. 12In this sense, herbs constitute the most significant resource for the research of therapeutically effective drugs.Consequently, the use of herbs in aromatherapy dates to around 1200 BC and these have been used in ancient civilizations for medical purposes, among others. 14,15cording to National Center for Complementary and Integrative Health (NCCIH), Americans spend more than $30.2 billion per year on aromatherapy, as well as global spending is expected to grow to $5 trillion by 2050; showing how this approach has gained acceptance and popularity recently.Aromatherapy uses essential oils (EOs) to cause a positive effect on mood, behavior, and wellness, in fact, several investigations have evaluated the therapeutic effects of EOs in mental health.However, further studies are needed to support the use of EOs in the treatment of mental health disorders.Therefore, the aim of this study was to evaluate the effect of Rosmarinus officinalis essential oil on anxiety, depression, and sleep quality.

Plant material
Fresh leaves of Rosmarinus officinalis were obtained from a local market and verified by Segundo Leiva Gonzales, Biol, and deposited at the Herbarium Antenor Orrego (HAO) of Antenor Orrego University.

Essential oils extraction
The leaves of Rosmarinus officinalis were washed with distilled water to remove dust.The samples

Introduction:
The objective of this investigation was to evaluate the effect of Rosmarinus officinalis essential oil on anxiety, depression, and sleep quality.Methods: The essential oils were extracted by hydro-distillation using a modified Clevenger-type apparatus and the chemical composition was performed by Gas chromatography with flame ionization detection (GC-FID) and Gas chromatography mass spectrometry (GC-MS).Furthermore, a quasi-experimental study was conducted, where 81 participants were divided into two groups, comprising a waiting list control group and an experimental group treated with essential oils.Anxiety and depression indexes were evaluated using the Zung Self-Rating Anxiety Scale (SAS) and the Zung Self-Rating Depression Scale (SDS), and sleep quality was measured with the Pittsburgh Sleep Quality Inventory (PSQI).Results: Chemical analysis showed that 1,8 cineole (37.6%), α-pinene (19.9%), and linalool (15.8%) were the main components.Anxiety, depression, and sleep quality scores showed a decrease in the post-test study phase compared to the pre-test in the experimental group (p<0.05).A large size effect was found in the case of anxiety (d = were then dried using a forced air circulation stove at a temperature of 40 ° C for 24 h.Next, the samples were milled, and the powdered plant materials (100 g) were placed in a round bottom flask with 1000 ml of distilled water and connected to a modified Clevenger-type apparatus.Hydrodistillation was completed for 4 h after boiling.Finally, the oil was dried with anhydrous sodium sulphate (Na 2 SO 4 ) and stored in a refrigerator in amber glass vials at 4°C for further use in experiments. 18

Study design and sample
This was a quasi-experimental study, using a pre-test and post-test design with an experimental group and a waiting list control group.81 participants were divided into two groups, the first one treated with aromatherapy based on essential oil from Rosmarinus officinalis leaves (EG) and a waiting-list (WL) control group.

Instruments
Zung Self-Rating Anxiety Scale (SAS): This scale consists of 20 items, scored each one from 1 to 4 (1= non or a little of the time, 2 = some of the time, 3 = good part of the time, 4 = most of the time). 20This instrument shows the validity and reliability for the population and context applied. 21ng Self-Rating Depression Scale (SDS): This scale consists of 20 items.Each item is scored from 1 to 4 (1= none or a little of the time, 2 = some of the time, 3 = good part of the time, 4 = most of the time). 22his instrument shows the validity and reliability for the population and context applied. 23ttsburgh Sleep Quality Inventory (PSQI): This questionnaire contains 18 items which are grouped into seven components.The score of each item ranges from 0 to 3. The sum score of these seven components is considered as the total PSQI score which is 0-21.Scores greater than 5 show low sleep quality. 24For the present study, the validity and reliability test for the local population and context was determined using the item test method, finding values greater than 0.42 for each item; additionally, the reliability coefficient of 0.96 was found using the split-half method.

Study procedure
A free aromatherapy course was offered to students at a private university.The course was conducted completely online, because of Peruvian government restrictions for the global pandemic.87 undergraduate students were enrolled, and 81 took part in this research between March and April 2022.Inclusion criteria included students enrolled in the academic semester 2022-01.Meanwhile, exclusion criteria were participants with previous practice of meditation, tai chi or yoga, psychiatric or pharmacological treatment, pregnancy, flu, and loss of smell for COVID-19.The students belonged to two sections (A and B) of the same course.These sections were formed before the researchers' arrival; for this reason, there was no randomization.Section A was selected as EG and Section B as WL.After group selection, instruments were administered (pretest).Additionally, each participant received a weekly aromatherapy kit with everything they needed for oils applications, including an instruction manual.Each kit was sent each week to each student until the investigation was completed.In addition, online meetings were held weekly to monitor the applications and provide feedback.The EOs application was according to the methodology of Reza et al., 25 where the participants were asked to pour two drops of EOs on a cotton ball using a dropper.Next, the cotton ball was held under the participant's nose while closed his or her eyes and took 10 deep breaths.Next, the cotton was pinned to the collar of the participant for 30 minutes.After that, the participant unpinned and disposed of the cotton ball.WL control group did not receive any intervention until the end of the application of EG.The applications were every day for 4 weeks.At the end of intervention, the instruments were administered again (posttest).(Figure 1).All participants were informed about the investigation program goals and signed a consent form in which confidentiality and anonymity were guaranteed.The study protocol was approved by Institutional Review Board (IRB).Furthermore, this investigation was carried out in accordance with the Declaration of Helsinki.

Data analysis
Data were presented as mean ± standard deviation (SD).Differences in sociodemographic and clinical data of participants were analyzed using the Pearson Chi-Square and Likelihood-ratio tests.The Mann-Whitney U test was used to determine significant differences between the groups, while the Wilcoxon test was used to determine significant differences between the study phases, in which p < 0.05 was considered statistically significant.These tests were used because the data did not conform to the normal distribution.Cohen's D and Hedges' G were calculated between the groups for post-test scores, as well as statistical power.Statistical analysis was performed using SPSS v.27.0 (IBM Corp., Armonk, NY, USA).
Table 3 shows the mean score and SDs for stress, sleep quality, anxiety, and depression where all groups did not show differences in pretest scores (p> 0.05).However, in posttest scores there were significant differences between WL and EG (p<0.05).All these outcomes were calculated by Mann Whitney U test between groups.Besides, regarding to the study phase, WL scores did not show differences between pretest and posttest (p>0.05).Nevertheless, all variables showed differences between study phases (p<0.05).All these results were calculated using the Wilcoxon test.
Besides, Table 4 shows the scores obtained by the Cohen's D and Hedges' G test, as well as the statistical power (1-β); where values between 0.5 and 0.8 show moderate changes, as is the case of stress (d = 790; g = 0.789), sleep quality (d = 0.586; g = 0.588) and depression (d = 0.581; g = 0.582).In addition, a large size effect was found for anxiety (d = 1.491; g = 1.492).Finally, this table shows the observed power (1-β), where anxiety is the only variable with an adequate statistical power (1-β=0.996).It is noteworthy to specify that the standard for the desired power is 0.80.

DISCUSSION
The COVID-19 pandemic is considered a new stressor for mental health, therefore interventions in this area will be a priority in the next times. 26In this context, chemical compounds from essential oils used in aromatherapy produce subtle neurological and behavioral changes, helping to improve mood states and associated disorders. 17 this investigation, the main components were1,8-Cineole, α-pinene, and linalool, which is in accordance with research performed in Pakistan. 19Somehow, another research found the main components were piperitone, linalool and α-pinene. 29At the same time, other investigations also found these compounds among the majoritarian compounds but in different ranges and quantities. 28Therefore, the components found in our work are frequently present in rosemary Eos; 29 but the variations are due to phenological stages, that is the ontogenesis process related to morphological modifications in the structure of plant organs which influence in EOs content 30 , as well as environmental factors such as altitude, soil conditions and seasonality 31 ; in addition to genotype, extraction and drying methods. 28,32 the other hand, with respect to sociodemographic data of participants, the majority were young unmarried and female students, which is in accordance with the statistical information presented in  Peruvian universities and the Latin American context. 33,34In addition, an investigation affirms that university students have a good attitude towards CAM, especially herbal medicine. 35However, this point constitutes one of the limitations because it would be needed a wider age range and sample to generalize the outcomes.
Rosmarinus officinalis EOs evidenced in animal experimental models, a potential to reduce anxiety and depression. 36In addition, there is information that shows a positive effect in reducing stress and anxiety in experimental models with volunteers. 37All these data are in accordance with the outcomes found in this investigation, where anxiety and depression scores decreased in the posttest phase only in EG.Indeed, rosemary EOs showed a large size effect in anxiety and a medium size effect in depression.Thus, it is more effective on anxiety, which is verified with the adequate statistical power observed for this variable, conferring the relevance of these results, which may be explained by the chemical constituents as monoterpenes presented in rosemary EOs, one of them is 1,8-Cineole which evidenced anxiolytic effects in humans. 38Thus, some authors have suggested that 1,8-cineole may interact with the benzodiazepine site on the GABA A receptor, producing anxiolysis, however, this compound did not entirely agonize GABA A site, which means that other receptors could be involved. 39esides, α-pinene exerts anxiolytic activity due to its hypnotic action to modulate GABA A -BZD receptors. 40In addition, linalool, another monoterpene found in rosemary EOs is considered a mood stabilizer, which shows anxiolytic properties, and its inhalation may reduce salivary cortisol, implicated in stress, a common trigger for anxiety, however linalool may block stress and diminish anxiety due to its capacity to restore catecholamine imbalance as well to reduce stressinduced ACTH levels. 41 the case of depression, monoterpenes such as linalool and β-pinene exert action through their interaction with the serotonergic route through postsynaptic 5-HT 1A receptors and the adrenergic system through α2-receptors, which play an important role in stress-induced behavioral changes. 42In fact, monoterpenes can be inhaled and affect directly the central nervous system, interacting with dopaminergic receptors such as D1 receptors, which the mechanism followed by most of the antidepressant drugs. 43Nevertheless, an investigation found that a single intraperitoneal dose of 1-8 cineole did not show antidepressant effects, however, most antidepressants in the market need an average of 2 to 4 weeks of administration to get therapeutic effects, 44 in addition, a potential pathway to EOs is a direct penetration of their molecules to brain areas via olfactory nerve, causing a rapid effect. 45garding to sleep, EG after treatment with EOs had significantly better sleep quality than CG, establishing a medium size effect and less power than the minimal required, which means that can rosemary EOs may improve moderately sleep quality, but the outcomes cannot be generalized.In this sense, there is evidence that rosemary extracts may improve sleep quality, 46 but human data related to these EOs is limited, however, components such as α-pinene displayed a sleep-increasing behavior by direct binding to GABA A -benzodiazepine (GABA A -BZD) receptors modulating partially BZD binding site, causing an hypnotic effect; in fact, this monoterpene may reduce sleep latency and enhanced non-rapid eye movement sleep (NREMS), which helps the body wind down as well as fall into a deep sleep, all these without affecting the rapid eye movement sleep (REMS) and delta action. 47Moreover; 1,8-cineole may also present the effect in question, possibly through the modulation of dopaminergic and glutamatergic systems. 48In addition, an investigation also found that linalool acupoint application therapy improved the sleep rate, sleep latency, and sleep duration. 49In fact, the sedative effect of linalool is dose dependent and includes the hypnotic effect, thus the most feasible hypothesis is that linalool acts as an antagonist of the excitatory neurotransmitter glutamate binding to glutamatergic N-methyl-D-aspartate (NMDA) receptors, which play a critical role in diurnal rhythmicity, sleep, and memory consolidation. 50nally, these results are not definitive because of limitations, thus future studies should be elucidated to generalize the results and specify the mechanism of action as well to explain if the effects are due to a particular compound or to the synergism of all.

CONCLUSION
The essential oils extracted from Rosmarinus officinalis are more effective for anxiety and show moderate changes in depression and sleep quality.

Figure 1 :
Figure 1: Flowchart of the study.

Table 2 : Sociodemographic and clinical data of participants.
a p-value is calculated by Pearson Chi-Square test.b p-value is calculated by Likelihood-ratio test.