“Burning Mouth Syndrome” (BMS) is charac- terized by burning sensation and pain in the mouth with or without inflammatory signs and specific lesions1. Synonyms found in scientific literature include glossodynia, oral dysesthesia, glossopyrosis, glossalgia, stomatopyrosis and stomatodynia1-4. It usually affects women aged between 40 and 60 years and the prevalence in the general population is 3.7%; BMS affects women in 65% of reported cases5. BMS general- ly shows three main aspects: mouth pain, alter- ation in taste and referred xerostomia in absence of visible mucosal lesions in the mouth. Pain is from moderate to severe in burning sensation, mainly affecting the dorsum and the tongue tip and it may persist along years. Pain may also be present in the gums, lips and jugal mucosa, with no visible lesions following pharyngeal inspec- tion. Pain increases, day by day, in states of anxi- ety, fatigue, excessive speaking and when ingest- ing hot and seasoned food; pain subsides with cold foods, work and recreation. Burning in the mouth does not manifest clinically in the zones where the peripheral nerves branch off, in fact, it typically affects more than one site. The etiology of BMS is really difficult to assess; in fact, there may be more than one etiological factors. Pa- tients seek help from a variety of medical spe- cialists, including dentists and dermatologists and they often try a range of therapies, such as: corticosteroids, analgesics, antibiotics, estrogens, retinoids and psychotropic drugs. However, none of the listed treatments were found to be really effective.
Hakeberg et al1, focusing on psychological as- pects of women with BMS, observed that all pa- tients in their study had gone through situations of great stress or disappointment in their lives, culminating with the appearance of mouth pain.
Furthermore, a recent article proposes an hy- pothesis focused on a neurodegenerative cause of BMS: chronic anxiety or post traumatic stress, as- sociated to menopause, leads to a dysregulation of the adrenal production of steroids7. One conse- quence is a decreased or modified production of some major precursors for the neuroactive steroid synthesis occurring in the skin, mucosa and ner- vous system. This results in neurodegenerative al- terations of small nerves fibers of the oral mucosa and /or some brain areas involved in oral somatic sensations. These neuropathic changes become ir- reversible and precipitate the burning pain, dys- geusia and xerostomia associated with stomatody- nia, all of which involve thin nerve fibers8.
The aim of the present study is to assess the re- lationship between Burning Mouth Syndrome (BMS) and oxidative stress in female patients, supposing that stomatodynia may be caused by an imbalance between Reactive Chemical Species (RCS) production and their elimination. In fact, oxidative stress represents an imbalance between the production and manifestation of reactive oxy- gen species (ROS) and a biological system’s abili- ty to readily detoxify the reactive intermediates or to repair the resulting damage. Disturbances in the normal redox state of tissues can cause toxic ef- fects through the production of peroxides and free radicals that damage all components of the cell, including proteins, lipids, and DNA. Some ROS can even act as messengers through a phenome- non called redox signaling. In the scientific litera- ture, oxidative stress has been associated with sev- eral diseases, such as neurodegenerative diseases9 or psychic impairments, such as ADHD (attention deficit hyperactivity disease)10 and anxiety11.

Materials and Methods

Eighteen female patients, aged between 54 and
68 years old, with diagnosis of primary Burning M o u t h S y n d r o m e i n a c c o r d a n c e w i t h t h e “Headache Classification Committee of the Inter- national Headache Society (2004) criteria”12, were evaluated between April 2009 and January
2011. All patients had primary BMS for more than 2 years and less than 5 years, and had no oral infections or other lesions and no diseases belonging to the exclusion criteria. A visual-ana- logue scale (V.A.S.)13 was used in order to evalu- ate the intensity pain referred by patients during the first clinical evaluation and the subsequent follow-up (Table I).
Exclusion criteria included Sjögren syndrome, rheumatological diseases, diabetes, hyper- or hy- p o t h y r o i d i s m , g e n e r a l i z e d p a i n , h i s t o r y o f surgery in the facial/oral region, Trigeminal Glossopharyngeal or Vagus neuralgia and smok- ing habit; moreover, we excluded all patients suf- fering from other disorders and clinical condi- tions which could have caused “oxidative stress”. Informed written consent was obtained from all patients recruited for this study.
Oxidative stress assessment was performed by means of an integrated analytical system com- posed of a photometer and a mini-centrifuge (FRAS4, H&D s.r.l., Parma, Italy). Samples of whole capillary blood were taken by a finger puncture in a heparinized tube and immediately centrifuged; a small amount of samples plasma (10 µL) were thereafter tested for total oxidant capacity (d-ROMs test) and biological antioxi- dant potential as iron-reducing activity (BAP test) (Diacron International s.r.l., Grosseto, Italy). The d-ROMs test is based on the ability of a plas- ma sample to oxidise the N,Ndiethylparaphe- nilendiamine (not coloured) to its radical cation (red colored); the reaction is monitored photometrically at 37°C at 505 nm and the results are ex p r e s s e d a s “ C a r r a t e l l i U n i t s ” ( C A R R U ,
∆Abs5050 nm/min). The oxidant capacity of plasma against N, N-diethylparaphenilendiamine is mainly due to hydroperoxides, with the contri- bution of (ferr) oxidase activity of ceruloplasmin and mieloperoxidase.

Normal Values of d-ROMs Test Ranges
Between 250 and 300 CARR Units
The BAP test is based on the ability of a plas-
ma sample to reduce the iron of a coloured com- plex containing ferric ions to its ferrous deriva- tive, not coloured; the reaction is monitored pho- tometrically at 37°C at 505 nm and the results are expressed in mol/L of reduced iron (using ascorbic acid as standard reference). Such bio- logical antioxidant potential is mainly due to vit- amin C, uric acid, bilirubin, albumin and toco- pherols hydroperoxides. Normal values of BAP test are assessed over 2200 µM.
BAP test values over 2200 µM were detected on the same machine using the same lot of kits; all tests were performed by the same operator and the analytical instrumentation was calibrated before the analytical session by means of plasma with known values of d-ROMs and BAP test. In order to further check the reliability of the analy- sis mean, we performed the same tests on sam- ples of whole capillary blood taken from 5 healthy volunteers and we had values of d-ROMs and BAP test just like we expected to find in healthy subjects; our 5 healthy volunteers were 5: these Caucasian females were aged between 60 and 67 years old.

This Study Was Conducted in Two
Different Times for Each Patients
First time (T0)s: it was performed in the same day in which each individual patient came to one of the Authors, referring about her symptomatol-
ogy consisting of stomatodynia and unbearable pain (according to VAS reported in Table I); a sample of capillary blood was then taken up and subjected to BAP-test (normal values > 2200
µM) and d-ROMs test (normal values between
250 and 300 U CARR).
Second time (T1): it was performed among 7 and 10 days after the latest manifestation of pain. All data were expressed as mean value ± SD.

Statistical Analysis
T0 data were statistically compared to T1 data performing the “paired t-test”.

dROMs
Paired t test
t = 6.673; df = 17; Two-tailed p <0.001
Number of pairs 18
Mean of differences 65.11
95% confidence interval 44.52 to 85.70
R squared 0.7237
Correlation coefficient (r) 0.2269

BAP
Paired t test
t = 8.655; df = 17; Two-tailed p < 0.001
Number of pairs 18
Mean of differences -512.0
95% confidence interval -636.8 to -387.2
R squared 0.8150
Correlation coefficient (r) 0.09435

Results
On recruitment, the mean value of d-ROMs test data resulted 429 ± 65, while the mean value of BAP Test data resulted 1880 ± 154.
The first follow-up visit at 7 days after the lat- est manifestation of pain (T1) revealed an average increase in BAP test of 14 ± 5% and an average reduction in d-ROM test of 28 ± 9%.
The ANOVA test showed high statistical sig- nificance (p < 0.001) between compared data.
The present study clearly demonstrated a sig- nificant correlation between “oxidative stress” and “Burning Mouth Syndrome” in female pa- tients.

Discussion
The aim of the present study was to investigate about a possible correlation between the “Burn- ing Mouth Syndrome” and oxidative stress. Up to now, no study, in the scientific literature, was performed in this light.
The analysis of results indicates that female patients, affected by Burning Mouth Syndrome, show significantly different d-ROMs and BAP levels, compatible with an oxidative stress condi- tion, if compared to the general population. Moreover, after the most painful phase of BMS, the d-ROMs and BAP levels progressively return to normal, even if the oxidative stress appears still significantly higher 7 days after this first phase, compared to the normal values, as well as seen by our research team. We considered even
the possibility of an increasing of oxidative stress levels as direct consequence to the sensation of chronic pain affecting the patients with BMS.
Although the increased ROS and the consequen- tial involvement of oxidative stress have been re- garded as important mechanisms in the pathogene- sis of pain, by reducing GABA inhibitory influence on SG neurons that are involved in pain transmission, the stability of measurements of biomarkers of oxidative stress in blood should remain valuable
up to 36-48h after the active phase of oxidative damage15. However, this study clearly showed d- ROMs and BAP levels still significantly higher 7 days after this active phase of oxidative damage, compared to the normal values.

Conclusions

The present study underlines the high levels of oxidative stress in female patients affected by Burning Mouth Syndrome. This investigation confirms the importance and the effectiveness of immediate antioxidant treatments in the BMS in order to prevent or decrease the onset of oxida- tive stress and the consequent increased risk of oxidative-related systemic diseases. Due to the importance of this preliminary result, we are con- scious to proceed in evaluating a much more number of patients, considering this study only as a pilot study, devoted to let people know, for the first time, the suppos ed and concretely demonstrated relationships.

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AUTHORS
M. TATULLO, M. MARRELLI, S. SCACCO, M. LORUSSO, S. D’ORIA, R. SABATINI, A. ABBINANTE, R. CAGIANO, F. INCHINGOLO
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