Saliva Samples Research Articles

Catalytic competence of gold clusters decorated yellow 2 G polymer composite film for voltammetric sensing of dopamine and nicotine in biological fluids

A novel, simple, cost-effective, easy-to-prepare, and timesaving electrochemical sensor based on gold clusters decorated yellow 2 G polymer composite film was fabricated and comprehensively characterized for the determination of dopamine and nicotine in real samples. When gold clusters were modified on the yellow 2 G polymer film, a signal increase of 204 % for dopamine and 526 % for nicotine was obtained compared to the activated bare pencil graphite electrode. The signals at 90 mV for dopamine and 780 mV for nicotine were utilized as analytical responses. The fabricated platform serves noteworthy detection sensitivity values of 79.02μA.μM−1.cm−2 and 14.61μA.μM−1.cm−2 with a notable limit of detection values of 58.8 nM and 7.2 nM for dopamine and nicotine, respectively. The electrochemical method has linear ranges of 0.075–5 µM and 6–10 µM for dopamine alone, 0.01–10 µM and 12–25 µM for nicotine alone, 0.01–10 µM and 12–50 µM for nicotine in the presence of dopamine, 0.2–10 µM and 12–60 µM for dopamine in the presence of nicotine, and 0.4–2, 2–10 and 12–90 µM for dopamine and 0.2–8, 10–20 and 25–90 µM for nicotine at the increasing amounts of each substance. Moving forward, the developed method exhibits splendid accuracy and selectivity for determining dopamine and nicotine in biological samples of human saliva, serum, and urine.

Aggregation-Induced Emission Luminogen: Role in Biopsy for Precision Medicine.

Biopsy, including tissue and liquid biopsy, offers comprehensive and real-time physiological and pathological information for disease detection, diagnosis, and monitoring. Fluorescent probes are frequently selected to obtain adequate information on pathological processes in a rapid and minimally invasive manner based on their advantages for biopsy. However, conventional fluorescent probes have been found to show aggregation-caused quenching (ACQ) properties, impeding greater progresses in this area. Since the discovery of aggregation-induced emission luminogen (AIEgen) have promoted rapid advancements in molecular bionanomaterials owing to their unique properties, including high quantum yield (QY) and signal-to-noise ratio (SNR), etc. This review seeks to present the latest advances in AIEgen-based biofluorescent probes for biopsy in real or artificial samples, and also the key properties of these AIE probes. This review is divided into: (i) tissue biopsy based on smart AIEgens, (ii) blood sample biopsy based on smart AIEgens, (iii) urine sample biopsy based on smart AIEgens, (iv) saliva sample biopsy based on smart AIEgens, (v) biopsy of other liquid samples based on smart AIEgens, and (vi) perspectives and conclusion. This review could provide additional guidance to motivate interest and bolster more innovative ideas for further exploring the applications of various smart AIEgens in precision medicine.

Freeze-dried wafers containing sesame oil for alleviation of dry mouth

A dry mouth is a condition with significant morbidity. Conventional saliva substitutes are expensive, short-lived, and prone to contamination. This study presents a Freeze-Dried Sesame Oil Emulsion Saliva Substitute (FD-SOESS), which combines the advantages of sesame oil through a freeze-drying process and assesses its effectiveness. High-speed homogenization (HSH) as well as high-pressure homogenization (HPH) techniques were employed to fabricate the FD-SOESS. The formulations that underwent the additional HPH step demonstrated improved physical properties, such as lower viscosity and faster redispersion times, compared to HSH alone. The optimized formulation (F11) possessed a near neutral pH of 6.67 and rapid re-dispersibility that was <141.5 s), which is acceptable for oral administration. Additionally, the FD-SOESS formulation revealed heterogeneous porous structures with a layered arrangement of different pore sizes, and the resulting formulation had low moisture content (2.76 %). The formulation met the microbial limit test. The antioxidant IC₅₀ of sesamin was found to be 0.318 μM. Furthermore, it was not toxic to periodontal cells at 50 μg/mL or 100 μg/mL. Human periodontal cell lines subjected to scratch and treated with FD-SOESS demonstrated notable migration that achieved complete wound closure within two days. The administration of FD-SOESS led to a notable elevation in the mucus concentrations found in the saliva samples of all participants. Overall, the results indicated that FD-SOESS is a highly promising alternative to saliva and may improve oral health for individuals suffering from dry mouth.

Estradiol and progesterone from pregnancy to postpartum: a longitudinal latent class analysis.

During the peripartum, women undergo significant hormonal changes that are crucial for fetal development and a healthy pregnancy and postpartum period for mother and infant. Although several studies have determined healthy norm ranges of estradiol and progesterone, there are discrepancies among the reports, rendering it unclear which hormone levels are linked to adverse health outcomes. To account for the impact of sex steroid patterns on health outcomes in mothers and children, a longitudinal assessment of different parameters is needed. We longitudinally assessed a cohort of 130 women over five months during pregnancy and postpartum. The women provided saliva samples and completed psychosocial questionnaires. Hormone analyses were conducted using enzyme-linked immunosorbent assay (ELISA). Different parameters of estradiol and progesterone were analyzed and evaluated in relation to psychometric variables. To examine the presence of heterogenous hormonal trajectories in the peripartum, we applied group-based trajectory modelling as a special case of latent-class group analysis. Estradiol and progesterone levels rose towards the end of pregnancy and dropped sharply after birth, with considerable individual variation, particularly during pregnancy. However, their ratio remained stable. We identified three estradiol trajectory subgroups and two progesterone subgroups. Age influenced progesterone levels, with older pregnant women having higher levels than younger women. Anxiety and depressive symptoms had a predictive value for trajectories of specific subgroups of women. The study also revealed two distinct subgroups regarding the course of estradiol and progesterone fluctuations as well as their ratio. This study provides insights into the course and fluctuation of salivary estradiol and progesterone levels among healthy women during the peripartum period, highlighting significant variations in hormone levels but stability in their ratio during this time. The finding of distinct sex steroid courses in the peripartum is new and suggests the need for further research to explore their impact on health outcomes. Our preliminary results suggest that hormonal fluctuations at the end of pregnancy appear to be a normal occurrence and might even be a protective factor for associated psychological symptoms and sleep disturbances in women.

Enantiospecific profiling of D-amino acid for gastric cancer diagnosis by using a biocatalytic MHOF nanoreactor

The dynamic variation of D-amino acids (D-AAs) in human saliva has great potential as the indicator for individual physiological states and disease progression; however, enantiospecific profiling of D-AAs is a great challenge. In this work, a nanoreactor based on mesoporous hydrogen-bonded organic framework (MHOF) has been fabricated for the enantiospecific profiling of D-AAs. The nanoreactor is prepared by using MHOFs as the scaffold to organize the biocatalytic cascade of D-AA oxidase (DAAO) and horseradish peroxidase (HRP). In the meanwhile, the large, stable, and robust transport channels of MHOFs allow the unimpeded transport of the involved biocatalytic substrates. Once D-AA substrates efficiently transport into MHOFs, the chromogenic reactions of 3,3′,5,5′-tetramethylbenzidine (TMB) and H2O2 can be activated. So, using this MHOF nanoreactor, enantiospecific and visual detection of D-AAs can be achieved. Moreover, the stable nanoreactor can be operated in complex solution and utilized for the profiling of D-AA levels in saliva samples collected from gastric cancer patients, suggesting that D-AAs may be a powerful fingerprint for the diagnosis of gastric cancer. Therefore, the fabricated MHOF-based nanoreactor may provide an emerging tool for the enantiospecific profiling of D-AAs and supports the D-AA fingerprint as the indicator of gastric cancer diagnosis.

Effect of a Novel Food Rich in Miraculin on the Oral Microbiome of Malnourished Oncologic Patients with Dysgeusia

Background/Objectives: Dysgeusia contributes to the derangement of nutritional status in patients with cancer as well as worsening the quality of life. There has been a lack of effective treatments for taste disorders provided by the pharmaceutical industry. Methods: This was a pilot randomized, parallel, triple-blind, and placebo-controlled intervention clinical trial in which 31 malnourished patients with cancer and dysgeusia receiving antineoplastic treatment were randomized into three arms [standard dose of DMB (150 mg DMB/tablet), high dose of DMB (300 mg DMB/tablet) or placebo (300 mg freeze-dried strawberry)] for three months. Patients consumed a DMB or placebo tablet before each main meal. Using the nanopore methodology, we analyzed the oral microbiome of patients with cancer using saliva samples. Results: All patients with cancer and dysgeusia had dysbiosis in terms of lower bacterial diversity and richness. DMB consumption was associated with changes in oral microbiome composition. Neither selected bacteria nor taste perception, type of diet, and cytokine levels were associated with mucositis. Likewise, alcohol and tobacco consumption as well as general and digestive toxicity due to systemic therapy were not associated with specific changes of the oral microbiome, according to logistic binary regression. The standard dose of DMB resulted in a lower abundance of Veillonella compared with the high DMB dose and placebo at 3 months after intervention with DMB. In particular, some species such as Streptococcus parasanguinis, Veillonella parvula, and Streptococcus mutans were less abundant in the DMB standard-dose group. Additionally, the consumption of a standard dose of DMB revealed a negative association between the concentrations of TNF-α and the abundance of species such as Streptococcus thermophilus, Streptococcus pneumoniae, Streptococcus dysgalactiae and Streptococcus agalactiae. Conclusions: Accordingly, regular DMB consumption could modify the oral microbiome in patients with cancer and dysgeusia, which may contribute to maintaining an appropriate immune response. However, as the present pilot study involved a small number of participants, further studies are necessary to draw robust conclusions from the data.

Salivary microbiota composition before and after use of proton pump inhibitors in patients with laryngopharyngeal reflux: a self-control study

BackgroundIssues associated with proton pump inhibitor (PPI) usage have been documented. PPIs affect the gastrointestinal microbiome, as well as the saliva microbiota of healthy individuals. However, the alterations in the saliva microbiota of laryngopharyngeal reflux (LPR) patients remain unclear. This study aims to examine the composition of saliva microbiota in LPR patients before and after PPI usage through a self-controlled study.MethodsThirty-two adult LPR patients participated in the study. Saliva samples were collected before and after an 8-week regimen of twice-daily administration of 20-mg esomeprazole. The impact of PPI administration on bacterial communities was assessed using 16 S rRNA gene sequencing. The functional and metabolic changes in saliva microbial communities after PPI usage were analyzed using PICRUSt2 based on our 16 S rRNA gene sequencing results.ResultsThe alpha diversity within the salivary microbiota, as measured by the PD-whole-tree index, exhibited a significant difference between samples collected before and after PPI application (P = 0.038). Additionally, PCoA analysis of unweighted UniFrac distances (beta diversity) revealed distinct separation of saliva sample microbiota structures before and after PPI application in LPR patients, with statistical significance (Adonis test, R2 = 0.063, P< 0.010). Taxon-based analysis indicated that PPI administration increased the abundance of Epsilonproteobacteria, Campylobacterales, Campylobacteraceae, Campylobacter, and Campylobacter_gracilis, while reducing the abundance of Lactobacillaceae and Lactobacillus in salivary samples ( P< 0.050). Using LEfSe to compare bacterial abundances, Bacillaceae and Anoxybacillus were found to be enriched before PPI usage in LPR patients. Furthermore, the proportion of genes responsible for indole alkaloid biosynthesis in the salivary microbiota of LPR patients significantly increased after PPI therapy (P< 0.050).ConclusionsThese findings indicate that PPIs induce alterations in the salivary microbiota of LPR patients.Chinese clinical trial registryNo. ChiCTR2300067507. Registered on January 10,2023 retrospectively.Level of evidence4.

Multi-site analysis of biosynthetic gene clusters from the periodontitis oral microbiome.

Background. Bacteria significantly influence human health and disease, with bacterial biosynthetic gene clusters (BGCs) being crucial in the microbiome-host and microbe-microbe interactions.Gap statement. Despite extensive research into BGCs within the human gut microbiome, their roles in the oral microbiome are less understood.Aim. This pilot study utilizes high-throughput shotgun metagenomic sequencing to examine the oral microbiota in different niches, particularly focusing on the association of BGCs with periodontitis.Methodology. We analysed saliva, subgingival plaque and supragingival plaque samples from periodontitis patients (n=23) and controls (n=16). DNA was extracted from these samples using standardized protocols. The high-throughput shotgun metagenomic sequencing was then performed to obtain comprehensive genetic information from the microbial communities present in the samples.Results. Our study identified 10 742 BGCs, with certain clusters being niche-specific. Notably, aryl polyenes and bacteriocins were the most prevalent BGCs identified. We discovered several 'novel' BGCs that are widely represented across various bacterial phyla and identified BGCs that had different distributions between periodontitis and control subjects. Our systematic approach unveiled the previously unexplored biosynthetic pathways that may be key players in periodontitis.Conclusions. Our research expands the current metagenomic knowledge of the oral microbiota in both healthy and periodontally diseased states. These findings highlight the presence of novel biosynthetic pathways in the oral cavity and suggest a complex network of host-microbe and microbe-microbe interactions, potentially influencing periodontal disease. The BGCs identified in this study pave the way for future investigations into the role of small-molecule-mediated interactions within the human oral microbiota and their impact on periodontitis.

Some Pro-inflammatory Cytokines Levels in Staphylococcal infection

Staphylococci is gram-positive, spherical cells, they are commonly distributed in the environment often part of normal human flora. Staph. aureus, Staph. saprophyticus, and Staph. epidermidis are the three most significant clinically important pathogens. Cytokines are small secreted proteins (&lt;40 kDa), which are produced by nearly every cell to regulate and affect immune response (96). Proinflammatory cytokines are proteins that have been associated with cell proliferation, differentiation, and changes in gene expression, and synthesis of matrix proteins important to cell growth and tissue repair, these proteins are often released in a cascade, where an initial release of TNF is followed first by IL-1 and then by IL-6. 110 clinical samples include 110 throat swap and 60 saliva samples were collected from tonsillitis patients in Kirkuk of both gender and different age groups. Sterile cotton swab were used for throat swab and sterile container for saliva collection. The results showed that Staphylococcus aureus scored highest percentage (59%), followed by Staph. hemolyticus (15%) compared to Staph. epidermidis that scored lowest percentage (2.4%). A sensitivity test was carried out for all bacterial isolates, Staphylococcus spp. showed high resistance to Cefotoxin (, Benzyle-pencillin (, Ampicillin-Sulbactam(, Oxacillin (, Ceftriaxone (, and Erythromycin(, while Staphylococcus spp. showed high sensitive to Linezolid (100%), Teicoplanin (100%), Vancomycin (100%), and Rifampicin (100%). Staph. aureus scored high mean levels at IL-6 (14.51±13.1 ng/L) compared with other Staphylococcus spp. (0.52±0.1 ng/L), while Staph. aureus scored (90.47±77.79 pg/ml) compared with other Staphylococcus spp. (113.0±79.9 pg/ml).

Salivary Antimicrobial Peptide in Patients With Dementia Before and After Clinical Oral Rehabilitation Programme: A Randomised Controlled Trial.

Emerging evidence suggests a link between salivary metabolite changes and neurodegenerative dementia, with antimicrobial peptides (AMPs) implicated in its pathogenesis. We investigated the effects of a clinical oral rehabilitation programme tailored for dementia patients on salivary flow rate, AMP levels and oral health-related quality of life (OHRQoL). Eligible patients were randomly assigned to either the experimental group (EG; n = 28) or the control group (CG; n = 27). Both groups received a leaflet on oral health. In addition, the EG received an oral care intervention that included individual lessons on oral muscle exercises and oral self-care practices. Saliva samples and OHRQoL data were collected at baseline and follow-up visits. Generalised estimating equation models were used to analyse the changes over time. At the 3-month follow-up, EG showed significantly lower histatin 5 (HTN-5) levels (β = -0.08; effect size [ES] = 0.72) than CG. At 6 months, EG exhibited improved salivary flow rate (β = 0.89; ES = 0.89) and OHRQoL (β = 6.99; ES = 1.31) compared to CG. Changes in salivary flow rate (β = 4.03), HTN-5 level (β = -0.78) and beta-defensin 2 level (BD-2) (β = -0.91) at 3 months predicted improved OHRQoL at 6 months (all p < 0.05). Our clinical oral rehabilitation programme reduced the level of salivary HTN-5, increased salivary flow rate and enhanced OHRQoL in dementia patients. Furthermore, changes in salivary flow rate, HTN-5 level and BD-2 level were associated with improvements in patients' OHRQoL.

A sensitive immunoassay for the quantitation of Pig-MAP in pig saliva samples

The purpose of the study is to develop a sensitive assay for the proper quantification of the acute phase protein Pig-MAP in pig saliva samples. A time-resolved immunofluorometric assay (TR-IFMA) was developed using two pig-MAP-specific monoclonal antibodies. The limit of detection of the assay was 4 ng/mL, enough to measure pig-MAP concentration in saliva. Precision was evaluated for saliva samples of low, medium and high concentration, with inter assay CV of 4–14 % and inter-assay CV of 8–20 %. The assay kept linearity under dilution and a method comparison study performed with serum samples showed good correlation with ELISA. Median Pig-MAP concentration in saliva from healthy animals was 19 ng/mL whereas in pigs with different inflammatory conditions was 11 times higher. In the same animals median pig-MAP serum concentrations were 0.72 mg/mL in the healthy group and 4.61 in the diseased group. The Spearman coefficient of correlation between Pig-MAP concentration in serum and saliva was of 0.72. A correlation was also observed between the salivary concentration of pig-MAP and other two acute phase proteins such as haptoglobin (r = 0.62) or C-reactive protein (r = 0.65). The concentration of Pig-MAP in saliva of pigs with severe respiratory disease decreased significantly from a median value of 128 ng/mL at the time of disease detection to 8 ng/mL after 1 day of antibiotic therapy. Studies performed show that pig-MAP is present in saliva and this specimen may be an alternative to serum for pig-MAP quantification.

Examining hypothalamic-pituitary-adrenal axis genetic variation on cortisol and alpha-amylase reactivity during an explicit negative-evaluative stress induction.

Prior work consistently links additive genetic variation in the hypothalamic-pituitary-adrenal (HPA) axis-a multilocus genetic profile score (MGPS)-to depression risk in the context of stress exposure. However, despite that HPA MGPS variants were selected based on evidence that they elevate cortisol reactivity, there are surprisingly few tests of whether an HPA MGPS elevates cortisol reactivity to lab-based stress. Similarly, despite neurobiological connections and coordination between the HPA axis and the sympathetic nervous system (e.g., in the paraventricular nucleus and locus coeruleus), no work has tested whether an HPA MGPS influences sympathetic nervous system reactivity to stress. We investigated whether an HPA MGPS moderates the relation between lab-based stress and: (1) HPA activity indexed by cortisol, and/or (2) sympathetic activity indexed by salivary alpha-amylase, sAA. Emerging adults (N=152; mean age=19.5, largest subsample 44.4% Black/African American) were randomly assigned to one of two Trier Social Stress Test variations, a non-evaluative control, or an explicitly negative-evaluative condition. Participants provided DNA and repeated saliva samples for sAA and cortisol. The HPA MGPS did not significantly moderate the relationship between stress condition and cortisol or sAA reactivity, respectively; moreover, post-hoc tests highlight that individual polymorphisms showed non-significant effects in opposite directions from each other, cancelling out in aggregate. Findings suggest that the HPA MGPS's associations with cortisol reactivity are not as straightforward as initially believed. We speculate that the relationships of some HPA variants to biomarker reactivity may vary between modest lab-based stressors and the explicit negative-evaluative induction used here.

Is there an association between salivary immune and microbial profile with dental health in systematically healthy children?

ObjectiveThis study aimed to characterize the inflammatory profile of systemically healthy children’s saliva and its association with clinical diagnoses of caries and gingival inflammation.Materials and methodsUnstimulated saliva was collected from 100 children before clinical dental examinations. The saliva samples were analyzed for total protein and specific inflammatory cytokines (IL-10, IL-8, IL-6, and TNFα) with Bradford and ELISA assays, respectively. Salivary bacteria were quantified using a quantitative real-time polymerase chain assay. The salivary values were then correlated with age, DMFT index, plaque index (PI), and gingival index (GI).ResultsThe mean age of the cohort was 8.08 ± 0.23 years with 49% females, the mean DMF of the cohort was 2.64 ± 0.31, the mean GI was 0.51 ± 0.06, and the mean PI was 1.33 ± 0.07. Significant correlations were found between PI with DMFT and GI. Children with DMFT > 2 had significantly higher levels of IL-8 compared with children with DMFT ≤ 2. IL-6 and TNFα were significantly higher among children with PI > 1 than among children with PI ≤ 1.ConclusionsSalivary cytokine were found to be associate with clinical parameters as DMFT and PI, thus may be a potential tool that reflects dental health status.Clinical relevanceThe presence of salivary cytokines in children may reflect evaluation of dental caries and oral inflammation.

Salivary assessment of the immune/inflammatory responses and oxidative stress in older adults vaccinated with CoronaVac or ChadOx-1

BackgroundAlthough important information concerning COVID-19 vaccination is available, the effects of the CoronaVac and ChadOx-1 vaccines on immunity and the redox balance in the upper airway mucosa of the aged population are not fully understood. Therefore, the aim of this study was to investigate the impacts of two doses of the CoronaVac or ChadOx-1 vaccine on immune/inflammatory responses and oxidative stress in the airway mucosa of older adults.MethodsSeventy-six older adults of both sexes, with a mean age of 75.1 ± 6.4 years, were separated according to vaccination status into the CoronaVac (n = 52) and ChadOx-1 (n = 24) groups. Saliva samples were collected before (pre) and 30 days after (post) the administration of the second dose of the CoronaVac or ChadOx-1 vaccine to assess the levels of antibodies (sIgA and IgG), antimicrobial peptides, cytokines, and oxidant/antioxidant agents.ResultsThe immunogenicity in the ChadOx-1 group was 37.5% for sIgA and 25% for IgG, while that in the CoronaVac group was 18.9% for sIgA and 13.2% for IgG. Intergroup analysis revealed that (1) lower levels of IFN-α, IFN-γ, and IL-10 and a greater IFN-γ/IL-10 ratio, in addition to a greater IL-6/IL-10 ratio, were found in both the pre- and postvaccination periods, and (2) lower levels of total sIgA, IL-12p70, IL-17A, TNF-α, and the IL-12p70/IL-10 ratio, in addition to higher levels of specific sIgA for SARS-CoV-2 antigens and lysozyme, were observed only in the postvaccination period in the ChadOx-1 group than in the CoronaVac group. Intragroup analysis revealed (1) a significant increase in the salivary levels of total peroxides in the postvaccination period compared to those in the prevaccination period in both volunteer groups; (2) a decrease in the levels of lysozyme and the ratio between total antioxidant capacity (TAC) and total peroxides in the postvaccination period in the CoronaVac group compared with those in the prevaccination period; and (3) decreases in the TNF-α, IL-6, and IL-12p70 levels, and the IL-12p70/IL-10 ratio in the ChadoX-1 group, as well as a higher lactoferrin concentration in the postvaccination period than in the prevaccination period. Several positive and negative correlations between the parameters assessed here were found.ConclusionsIn general, the ChadOx-1 group exhibited improvements in both immune/inflammatory responses and redox balance and greater immunogenicity than did the CoronaVac group.

Estimation of Periodontal Inflamed Surface Area by Salivary Lactate Dehydrogenase Level Using a Test Kit.

Background: Salivary lactate dehydrogenase (LD) levels are a feasible and useful parameter for screening periodontal diseases. The periodontal inflamed surface area (PISA) is useful to clinically assess periodontal diseases. However, PISA is difficult to calculate and PISA-compatible screening kits are required. We aimed to investigate the association between salivary LD levels, using a test kit, and PISA and PISA-Japanese and determine the feasibility and reliability of the salivary LD test kit for evaluation of periodontal status. Methods: This study included 110 patients (66.4% female, median and 25-75 percentiles of age were 66.5 and 53.0-75.0 years, respectively) who visited the Dental University Clinic in Japan. Resting saliva samples were collected from each participant and LD levels were evaluated in real time using a kit featuring an integer scale ranging from 1 to 10. PISA and PISA-Japanese were calculated using periodontal parameters. Results: The median salivary LD level was 4.0. The medians of PISA and PISA-Japanese were 46.9 and 61.0, respectively. Salivary LD levels were positively correlated with the bleeding on probing rate (r = 0.626, p < 0.001), PISA (r = 0.560, p < 0.001), and PISA-Japanese (r = 0.581, p < 0.001). Conclusions: Our results suggest that salivary LD levels assessed using the salivary LD kit showed a significantly positive correlation with PISA and PISA-Japanese. In addition, we developed the PISA estimation formula using salivary LD levels measured with a test kit, sex, and age.

Humic acid-coated paper: An affordable mixed-mode weak-cation exchanger for isolating cocaine, codeine, and methadone from saliva samples for their direct infusion mass spectrometric analysis.

The use of natural materials as precursors for synthesizing sorptive phases can reduce the environmental impact of the sample preparation step. In this article, two natural materials, i.e., cellulose and humic acids, are combined, resulting in a porous and flat sorptive phase that combines hydrophobic and cation exchange interactions. The simple preparation, with low reagent consumption, and the high production capacity make this process affordable. The performance of the humic acid-coated paper has been studied towards three basic drugs (cocaine, codeine, and methadone), and it has allowed the determination of the targets in saliva by direct infusion tandem mass spectrometry. The simultaneous extraction of many samples and the rapid analysis (2 min) provide a high sample throughput. The resulting method has been validated following the International Conference on Harmonization (ICH) M10 validation guide. Working at the optimum conditions, the linear range was established from 5 to 500 μg·L-1 and the detection limits were established at 1.5 μg·L-1 for all the analytes. At the LOQ, the inter-day precision and intra-day precision, expressed as relative standard deviation, were better than 13.6% and 17.9%, respectively. For low/mid/high quality controls, the inter-day precision and intra-day precision were better than 11.8% and 9.1%, respectively. The accuracy, calculated as relative recovery, was in the range of 83.8-106.1%. Several samples from patients under treatment with codeine-containing prescriptions were finally analyzed.

Metabolomic insights into idiopathic xerostomia: The central role of caffeine metabolism in salivary biochemistry

ObjectiveThis study aims to delineate the salivary metabolomic profile of patients with idiopathic xerostomia using untargeted metabolomics techniques, with the goal of addressing the lack of clear diagnostic markers and providing insights into the pathophysiological mechanisms of the condition. DesignIn this observational, cross-sectional study, saliva samples from 33 patients with idiopathic xerostomia and 34 healthy controls were analyzed using Ultra Performance Liquid Chromatography Quadrupole Time of Flight Mass Spectrometry (UPLC-QTOF MS). Metabolomic profiling was complemented by multivariate statistical analysis to differentiate between affected individuals and controls. ResultsMetabolomic analysis delineated a pronounced differentiation between patients with idiopathic xerostomia and healthy controls. A total of 195 metabolites displayed significant differential expression, each with a variable importance in projection (VIP) greater than 1 and a P-value less than 0.05. Pathway enrichment analysis, according to the Kyoto Encyclopedia of Genes and Genomes (KEGG), identified 22 metabolites that participated in 18 distinct metabolic pathways. Among these, the caffeine metabolism pathway, characterized by notable alterations in impact values (VIP, P-value, Log2-fold change, Rich factor), emerged as the most significantly disrupted, underscoring its potential role in the pathophysiology of idiopathic xerostomia (P = 0.0000395). ConclusionsThe salivary metabolomic profiling revealed distinct alterations in idiopathic xerostomia, with a significant reduction in caffeine metabolism pathways, underscoring potential neuropathic involvement. This study advances our understanding of the metabolic alterations in xerostomia, suggesting that salivary metabolomics may offer viable biomarkers for diagnosing and understanding the etiology of idiopathic xerostomia. Future research should focus on therapeutic targeting of these metabolic disturbances and evaluating their reversibility with treatment.

B-185 A Comparison Between Two Oral Sample Collection Devices on Preserving High-Quality Genomic DNA

Abstract Background High-quality genomic testing significantly enhances the knowledge on gene-disease association, and greatly benefits future biomedical research and precision medicine. However, widely used saliva sample collection presents challenges with high-quality and quantity of human DNA and high bacterial DNA ratio. In this study, we validated the performance of two commercially available oral collection devices for their ability to preserve high-quality genomic DNA long-term at room temperature. Methods Fifty-one buccal samples in total from twelve donors were collected using two kinds of commercially available oral sample collection devices (iSWAB®-DNA (Device 1) and OCD-100 (Device 2)). All of the collected samples were stored at ambient temperature for up to 1 year. Collected samples were extracted with QIAamp Blood Mini Kit (Qiagen) at baseline, 30 days, 60 days, and 1 year. All DNA yields were measured using a Nanodrop One Microvolume UV-Vis Spectrophotometer; TapeStation System was used for determining the integrity of DNA. The bacterial DNA ratio was estimated by quantitative PCR with primers targeting the 16S rRNA gene known to be present in prokaryotes but not eukaryotes. Results Device 1 had higher yield (Device 1: 18.59 ± 8.98; Device 2: 6.44 ± 4.62), lower bacterial DNA ratio (Device 1: 2.06 ± 1.93; Device 2: 3.79 ± 4.58), and higher overall DNA integrity (Device 1: 6.1 ± 0.31; Device 2: 5.42 ± 0.96) compared to Device 2. Device 1 demonstrated longer post-collection sample stability at ambient temperature compared with Device 2 regarding DNA yield, bacterial DNA ratio and DNA integrity. No significant changes in DNA yield, integrity, and bacterial DNA ratio were observed after one year of storage in Device 1. No significant changes in DNA yield, integrity, and bacterial DNA ratio were observed after 60 days of storage in Device 2. However, Device 2 showed a significant increase in DNA yield, bacterial DNA ratio, and DNA integrity comparing baseline to 1 year. Conclusions Device 1 preserves higher quantity and quality of DNA from oral samples at ambient temperature for up to 1 year. These advantages significantly enhance efficiency and cost-effectiveness in downstream applications such as genotyping and sequencing, making it a valuable tool in genomics research with a commendable success rate.

Retrospective analysis of SARS-CoV-2 RNA detection in pooled saliva samples: An effective cost saving method.

Retrospective analysis of SARS-CoV-2 RNA detection in pooled saliva samples: An effective cost saving method.

Effects of preservatives on the activities of salivary enzymes

ObjectivesTo investigate the effects of common preservatives used in oral health care products on the enzymatic activities of lysozyme, peroxidase, and α-amylase in-solution and on-hydroxyapatite surface phases. DesignThe preservatives used in this study were sodium benzoate, methylparaben, propylparaben, and benzalkonium chloride. Hen egg-white lysozyme, bovine lactoperoxidase, and α-amylase from Bacillus sp. served as sources of purified enzymes. Human unstimulated whole saliva was used as a source of salivary enzymes. Hydroxyapatite beads were used as the surface phase. The preservatives were incubated with purified enzymes or saliva samples in-solution or on-hydroxyapatite surface phases, respectively. Enzymatic activities of lysozyme, peroxidase, and α-amylase were measured by hydrolysis of fluorescein-labelled Micrococcus lysodeikticus, oxidation of fluorogenic 2′,7′-dichlorofluorescin, and hydrolysis of fluorogenic starch, respectively. ResultsThe effects of the preservatives on the enzymatic activities of lysozyme and peroxidase were more distinct in the saliva samples than purified substances, and in the in-solution phase than on-hydroxyapatite surface phase, and the opposite was true for α-amylase. The most significant result was apparent decrease in peroxidase activities caused by the parabens in the in-solution phase (P<0.05). Sodium benzoate and parabens inhibited lysozyme activity in the in-solution phase, but differently for the purified and salivary lysozymes. Parabens and benzalkonium chloride inhibited the enzymatic activity of α-amylase from Bacillus sp., not saliva samples, only on-hydroxyapatite surface (P<0.05). ConclusionsEach preservative affected the enzymatic activities of lysozyme, peroxidase, and α-amylase differently. Based on the effects on salivary enzymes, sodium benzoate or benzalkonium chloride was recommended as preservatives rather than parabens.