Efficient Combination Chemo-Sonodynamic Cancer Therapy Using Mitochondria-Targeting Sonosensitizer-Loaded Polysorbate-Based Micelles.

Sonodynamic therapy (SDT), utilizing ultrasound (US) and sonosensitizers, holds immense potential as a noninvasive and targeted treatment for a variety of deep-seated tumors. However, the clinical translation of SDT is hampered by several key limitations in sonosensitizers, especially their low aqueous stability and poor cellular uptake. In this study, non-ionic polysorbate (Tween 80, T80) was adopted to formulate effective nanocarriers for the safe and efficient delivery of sonosensitizers to cancer cells. Mitochondria-targeting triphenylphosphonium (TPP)-conjugated chlorin e6 (Ce6) sonosensitizer was loaded into T80-based micelles for efficient SDT. Pro-oxidant piperlongumine (PL) was co-encapsulated with TPP-conjugated Ce6 (T-Ce6) in T80 micelles to enable combination chemo-SDT. T80 micelles substantially enhanced the cellular internalization of T-Ce6. As a result, T80 micelles loaded with T-Ce6 and PL [T80(T-Ce6/PL)] significantly elevated intracellular reactive oxygen species (ROS) generation in MCF-7 human breast cancer cells upon US exposure. Moreover, T-Ce6 exhibited selective accumulation within the mitochondria, leading to efficient cell death under US irradiation. Importantly, T80(T-Ce6/PL) micelles caused cancer-specific cell death by selectively triggering apoptosis in cancer cells through PL. This study demonstrated the feasibility of using T80(T-Ce6/PL) micelles for efficient and cancer-specific combination chemo-SDT.

Ferumoxytol Nanoparticles Target Biofilms Causing Tooth Decay in the Human Mouth.

Severe tooth decay has been associated with iron deficiency anemia that disproportionally burdens susceptible populations. Current modalities are insufficient in severe cases where pathogenic dental biofilms rapidly accumulate, requiring new antibiofilm approaches. Here, we show that ferumoxytol, a Food and Drug Administration-approved nanoparticle formulation for treating iron deficiency, exerts an alternative therapeutic activity via the catalytic activation of hydrogen peroxide, which targets bacterial pathogens in biofilms and suppresses tooth enamel decay in an intraoral human disease model. Data reveal the potent antimicrobial specificity of ferumoxytol iron oxide nanoparticles (FerIONP) against biofilms harboring Streptococcus mutans via preferential binding that promotes bacterial killing through in situ free-radical generation. Further analysis indicates that the targeting mechanism involves interactions of FerIONP with pathogen-specific glucan-binding proteins, which have a minimal effect on commensal streptococci. In addition, we demonstrate that FerIONP can detect pathogenic biofilms on natural teeth via a facile colorimetric reaction. Our findings provide clinical evidence and the theranostic potential of catalytic nanoparticles as a targeted anti-infective nanomedicine.

Mechanical characterization of vesicles and cells: A review.

Vesicles perform many essential functions in all living organisms. They respond like a transducer to mechanical stress in converting the applied force into mechanical and biological responses. At the same time, both biochemical and biophysical signals influence the vesicular response in bearing mechanical loads. In recent years, liposomes, artificial lipid vesicles, have gained substantial attention from the pharmaceutical industry as a prospective drug carrier which can also serve as an artificial cell-mimetic system. The ability of these vesicles to enter through pores of even smaller size makes them ideal candidates for therapeutic agents to reach the infected sites effectively. Engineering of vesicles with desired mechanical properties that can encapsulate drugs and release as required is the prime challenge in this field. This requirement has led to the modifications of the composition of the bilayer membrane by adding cholesterol, sphingomyelin, etc. In this article, we review the manufacturing and characterization techniques of various artificial/synthetic vesicles. We particularly focus on the electric field-driven characterization techniques to determine different properties of vesicle and its membranes, such as bending rigidity, viscosity, capacitance, conductance, etc., which are indicators of their content and mobility. Similarities and differences between artificial vesicles, natural vesicles, and cells are highlighted throughout the manuscript since most of these artificial vesicles are intended for cell mimetic functions.

Stiffness measurement of nanosized liposomes using solid-state nanopore sensor with automated recapturing platform.

This paper describes a method to gauge the stiffness of nanosized liposomes - a nanoscale vesicle - using a custom-made recapture platform coupled to a solid-state nanopore sensor. The recapture platform electrically profiles a given liposome vesicle multiple times through automated reversal of the voltage polarity immediately following a translocation instance to re-translocate the same analyte through the nanopore - provides better statistical insight at the molecular level by analyzing the same particle multiple times compared to conventional nanopore platforms. The capture frequency depends on the applied voltage with lower voltages (i.e., 100 mV) permitting higher recapture instances than at higher voltages (>200 mV) since the probability of particles exiting the nanopore capture radius increases with voltage. The shape deformation was inferred by comparing the normalized relative current blockade ( ΔI/I0̂) at the two voltage polarities to that of a rigid particle, i.e., polystyrene beads. We found that liposomes deform to adopt a prolate shape at higher voltages. This platform can be further applied to investigate the stiffness of other types of soft matters, e.g., virus, exosomes, endosomes, and accelerate the potential studies in pharmaceutics for increasing the drug packing and unpacking mechanism by controlling the stiffness of the drug vesicles.

Dried Saliva Spot (DSS) as a Convenient and Reliable Sampling for Bioanalysis: An Application for the Diagnosis of Diabetes Mellitus.

This paper proposes the dried saliva spot (DSS) as a convenient sampling technique for bioanalysis. The analytical method with the DSS was used for the determination of D,L-lactic acid (D,L-LA) and the D/L ratio of diabetic patients and prediabetic persons for the simple screening of the disease. The D,L-LA in the DSS was labeled with a chiral reagent (DMT-3(S)-Apy) for carboxylic acids and determined by UPLC-ESI-MS/MS. The limits of detection (signal-to-noise ratio (S/N) = 3) for the DSS analysis were on the amol level (∼30 amol). Because good stability, recovery, accuracy, and precision of the D,L-LA for the DSS method was also obtained from the proposed procedure, the DSS method was applied to the determination of the D- and L-isomers of LA of diabetic patients, and prediabetic and healthy persons. The D/L-LA ratio by the present DSS method and the HbA1c value in blood were well-correlated to the serious diabetic patients, whereas the relation in the prediabetic persons was not very good. The reason seems to be due to the rough saliva sampling, and not to the DSS method, because strict regulation was not requested for the prediabetic and healthy persons. In order to have a successful DSS analysis, the stability of the target molecule, the detection sensitivity to the target molecule, and the validated determination method are important.

Principal component analysis of molecularly based signals from infant formula contaminations using LC-MS and NMR in foodomics.

BACKGROUND: The challenge in developing analytical assessment of unexpected excess contaminations in infant formula has been the most significant project to address the widespread issue of food safety and security. Foodomics based on metabolomics techniques provides powerful tools for the detection of tampering cases with intentional contaminations. However, the safety and risk assessments of infant formula to reveal not only the targeted presence of toxic chemicals, but also molecular changes involving unexpected contaminations, have not been reported. In this study, a huge amount of raw molecularly based signals from infant formula was analysed using reversed phase and hydrophilic interaction chromatography with time-of-flight MS (LC-MS) and (1) H nuclear magnetic resonance (NMR) and then processed by a principal component analysis (PCA). RESULTS: PCA plots visualised signature trends in the complex signal-data batches from each excess contamination of detectable chemicals by LC-MS and NMR. These trends in the different batches from a portion of excess chemical contaminations such as pesticides, melamine and heavy metals and out-of-date products can be visualised from spectrally discriminated infant formula samples. CONCLUSION: PCA plots provide possible attempts to maximise the covariance between the stable lot-to-lot uniformity and excess exogenous contaminations and/or degradation to discriminate against the molecularly based signals from infant formulas. © 2015 Society of Chemical Industry.

Low aspect ratio micropores for single-particle and single-cell analysis.

This paper describes microparticle and bacterial translocation studies using low aspect ratio solid-state micropores. Micropores, 5 µm in diameter, were fabricated in 200 nm thick free-standing silicon nitride membranes, resulting in pores with an extremely low aspect ratio, nominally 0.04. For microparticle translocation experiments, sulfonated polystyrene microparticles and magnetic microbeads in size range of 1-4 µm were used. Using the microparticle translocation characteristics, we find that particle translocations result in a change only in the pore's geometrical resistance while the access resistance remains constant. Furthermore, we demonstrate the ability of our micropore to probe high-resolution shape information of translocating analytes using concatenated magnetic microspheres. Distinct current drop peaks were observed for each microsphere of the multibead architecture. For bacterial translocation experiments, nonflagellated Escherichia coli (strain HCB 5) and wild type flagellated Salmonella typhimurium (strain SJW1103) were used. Distinct current signatures for the two bacteria were obtained and this difference in translocation behavior was attributed to different surface protein distributions on the bacteria. Our findings may help in developing low aspect ratio pores for high-resolution microparticle characterization and single-cell analysis.

Use of solid-state nanopores for sensing co-translocational deformation of nano-liposomes.

Membrane deformation of nano-vesicles is crucial in many cellular processes such as virus entry into the host cell, membrane fusion, and endo- and exocytosis; however, studying the deformation of sub-100 nm soft vesicles is very challenging using the conventional techniques. In this paper, we report detecting co-translocational deformation of individual 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) nano-liposomes using solid-state nanopores. Electrokinetic translocation through the nanopore caused the soft DOPC liposomes (85 nm diameter) to change their shape, which we attribute to the strong electric field strength and physical confinement inside the pore. The experiments were performed at varying transmembrane voltages and the deformation was observed to mount up with increasing applied voltage and followed an exponential trend. Numerical simulations were performed to simulate the concentrated electric field strength inside the nanopore and a field strength of 14 kV cm(-1) (at 600 mV applied voltage) was achieved at the pore center. The electric field strength inside the nanopore is much higher than the field strength known to cause deformation of 15-30 µm giant membrane vesicles. As a control, we also performed experiments with rigid polystyrene beads that did not show any deformation during translocation events, which further established our hypothesis of co-translocational deformation of liposomes. Our technique presents an innovative and high throughput means for investigating deformation behavior of soft nano-vesicles.

Profiling of chiral and achiral carboxylic acid metabolomics: synthesis and evaluation of triazine-type chiral derivatization reagents for carboxylic acids by LC-ESI-MS/MS and the application to saliva of healthy volunteers and diabetic patients.

Novel triazine-type chiral derivatization reagents, i.e., (S)-1-(4,6-dimethoxy-1,3,5-triazin-2-yl)pyrrolidin-3-amine (DMT-3(S)-Apy) and (S)-4,6-dimethoxy-N-(pyrrolidin-3-yl)-1,3,5-triazin-2-amine (DMT-1(S)-Apy), were developed for the highly sensitive and selective detection of chiral carboxylic acids by UPLC-MS/MS analysis. Among the synthesized reagents, DMT-3(S)-Apy was a more efficient chiral reagent for the enantiomeric separation of chiral carboxylic acids in terms of separation efficiency by reversed-phase chromatography and detection sensitivity by ESI-MS/MS. The DMT-3(S)-Apy was used for the determination of 13 carboxylic acids in human saliva of healthy volunteers and diabetic patients. Various biological carboxylic acids including chiral carboxylic acids, and mono- and di-carboxylic acids were clearly identified in the saliva of healthy persons and diabetic patients. The concentrations of carboxylic acids detected in the saliva of diabetic patients were relatively higher than those in the healthy persons. Furthermore, the concentration of D-lactic acid (LA) and the ratio of D/L-LA in the diabetic patients were significantly higher than those in the healthy persons. The low ratio of D/L-LA in healthy persons was also identified to be independent of age and sex. These results suggest that the determination of the D/L-LA ratio in saliva might be applicable for the diagnosis of diabetes. Based on these observations, DMT-3(S)-Apy seems to be a useful chiral derivatization reagent for the determination not only of chiral carboxylic acids but also achiral ones. In conclusion, the proposed method using DMT-3(S)-Apy is useful for the carboxylic acid metabolomics study of various specimens.

Evaluation of a series of prolylamidepyridines as the chiral derivatization reagents for enantioseparation of carboxylic acids by LC-ESI-MS/MS and the application to human saliva.

Mass spectrometry has become a popular analytical tool because of its high sensitivity and specificity. The use of a chiral derivatization reagent for the mass spectrometry (MS) detection seems to be efficient for the enantiomeric separation of racemates. However, the number of chiral reagents for the liquid chromatography (LC)-MS/MS analysis is very limited. According to these observations, we are currently in the process of developing novel labeling reagents for chiral molecules in MS/MS analysis. The derivatization reagent that is effective for enhancing not only the electrospray ionization-MS/MS sensitivity but also the reversed-phase LC resolution of carboxylic acid enantiomers should have a highly proton-affinitive moiety and an asymmetric structure near the reactive functional group. Furthermore, the resulting derivative has to provide a characteristic product ion suitable for the selected reaction monitoring. Based upon these considerations, a series of prolylamidepyridines ((S)-N-pyrrolidine-2-carboxylic acid N-(pyridine-2-yl)amide (PCP2), (S)-N-pyrrolidine-2-carboxylic acid N-(pyridine-3-yl)amide, and (S)-N-pyrrolidine-2-carboxylic acid N-(pyridine-4-yl)amide) was synthesized as ideal labeling reagents for the enantioseparation of chiral carboxylic acids and evaluated in terms of separation efficiency and detection sensitivity by ultra-performance LC (UPLC)-MS/MS. Among the synthesized reagents, PCP2 was the most efficient chiral derivatization reagent for the enantioseparation of carboxylic acid. The Rs values and the detection limits of the derivatives of non-steroidal anti-inflammatory drugs, which were selected as the representative carboxylic acids, were in the range of 2.52-6.07 and 49-260 amol, respectively. The sensitive detection of biological carboxylic acids (detection limits, 32-520 amol) was also carried out by the proposed method using PCP2 and UPLC-MS/MS. The PCP2 was applied to the determination of carboxylic acids in human saliva. Several biological carboxylic acids, such as lactic acid (LA), 3-hydroxybutylic acid, maric acid, succinic acid, α-ketoglutalic acid, and citric acid, were clearly identified in the saliva of healthy persons and diabetic patients. Furthermore, the ratio of D-LA in diabetic patients was higher than that in normal subjects. Judging from these results, PCP2 seems to be a useful chiral derivatization reagent for the determination not only of chiral, but also achiral, carboxylic acids in real samples.

Augmentation genioplasty using discarded bone fragments following proximal segment osteotomy of the ramus in intraoral vertical ramus osteotomy (IVRO).

BACKGROUND: Based on a three-dimensional (3D) orthognathic simulation, this technical report introduces a method for augmentation genioplasty using a proximal bone fragment of the mandible, which is typically discarded in intraoral vertical ramus osteotomy (IVRO). RESULTS: A 43-year-old female patient diagnosed with Class III malocclusion, presenting with a protruding mandible and long facial height, underwent surgical treatment. The surgical plan involved mandibular setback position using IVRO and augmentation genioplasty. The 3D orthognathic surgery including augmentation genioplasty simulation was performed. An excessively elongated proximal segment was sectioned following IVRO. The inferior part of the sectioned proximal bone fragment of the mandible was positioned to align with the requirements of advancement genioplasty. After ensuring that the placement of the fragment matched that of the simulated surgery, each bone fragment was fixed. At 1.5 years post-surgery, the grafted bone on the augmentation genioplasty was well maintained, with slight bone resorption. CONCLUSIONS: Augmentation genioplasty using the proximal bone fragment of the mandible, which is typically discarded in IVRO, reduces the surgical complications associated with chin osteotomy. When a secondary genioplasty is required, genioplasty with osteotomy, movement of the cut bone fragments, partial bone-shaving osteotomy, and additional bone grafting are viable options.

Drug-loaded adhesive microparticles for biofilm prevention on oral surfaces.

The oral cavity, a warm and moist environment, is prone to the proliferation of microorganisms like Candida albicans (C. albicans), which forms robust biofilms on biotic and abiotic surfaces, leading to challenging infections. These biofilms are resistant to conventional treatments due to their resilience against antimicrobials and immune responses. The dynamic nature of the oral cavity, including the salivary flow and varying surface properties, complicates the delivery of therapeutic agents. To address these challenges, we introduce dendritic microparticles engineered for enhanced adhesion to dental surfaces and effective delivery of antifungal agents and antibiofilm enzymes. These microparticles are fabricated using a water-in-oil-in-water emulsion process involving a blend of poly(lactic-co-glycolic acid) (PLGA) random copolymer (RCP) and PLGA-b-poly(ethylene glycol) (PLGA-b-PEG) block copolymer (BCP), resulting in particles with surface dendrites that exhibit strong adhesion to oral surfaces. Our study demonstrates the potential of these adhesive microparticles for oral applications. The adhesion tests on various oral surfaces, including dental resin, hydroxyapatite, tooth enamel, and mucosal tissues, reveal superior adhesion of these microparticles compared to conventional spherical ones. Furthermore, the release kinetics of nystatin from these microparticles show a sustained release pattern that can kill C. albicans. The biodegradation of these microparticles on tooth surfaces and their efficacy in preventing fungal biofilms have also been demonstrated. Our findings highlight the effectiveness of adhesive microparticles in delivering therapeutic agents within the oral cavity, offering a promising approach to combat biofilm-associated infections.

High-throughput LC-MS/MS based simultaneous determination of polyamines including N-acetylated forms in human saliva and the diagnostic approach to breast cancer patients.

The determination of polyamines and their N-acetylated forms was performed by ultraperformance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS). The polyamines efficiently reacted with 4-(N,N-dimethylaminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole (DBD-F) in 0.1 M borax (pH 9.3) at 60 °C for 30 min. The resulting derivatives were analyzed by electrospray ionization (ESI)-MS and sensitively detected by selected reaction monitoring (SRM). Furthermore, a rapid separation of the polyamine derivatives within 10 min was performed by UPLC using an antipressurized column packed with 1.7-µm octadecylsilyl (ODS) silica gel. The limits of detection (S/N = 3) on the SRM chromatograms were at the attomole level (9-43 amol). This procedure was used to successfully determine 11 polyamines, including their N-acetylated forms, in the saliva of patients with primary and relapsed breast cancer and healthy volunteers. The level of several polyamines (Ac-PUT, Ac-SPD, Ac-SPM, DAc-SPD, and DAc-SPM) increases in breast cancer patients. Furthermore, the levels of three polyamines (Ac-SPM, DAc-SPD, and DAc-SPM) were significantly higher only in the relapsed patients. The present method proved highly sensitive and is characterized by specificity and feasibility for sample analysis. Consequently, the proposed method is useful for the noninvasive salivary diagnosis of cancer patients and could be applied to determine polyamines in several specimens of biological nature.

A novel derivatization reagent possessing a bromoquinolinium structure for biological carboxylic acids in HPLC-ESI-MS/MS.

A novel bromoquinolinium reagent, i.e. 1-(3-aminopropyl)-3-bromoquinolinium bromide (APBQ), was synthesized for the analysis of carboxylic acids. A simple and practical precolumn derivatization procedure using the APBQ in RP chromatography and MS (HPLC-MS) has been developed using bile acids and free fatty acids, as the representative carboxylic acids in biological samples. The APBQ efficiently reacted with carboxylic acids at 60°C for 60 min in the presence of N,N-dicyclohexylcarbodiimide and pyridine as the activation reagents. Because the APBQ possesses a bromine atom in the structure, the identification of a series of carboxylic acids was easily achieved due to the characteristic bromine isotope pattern in the mass spectra. The APBQ also has a quaternary amine structure, thus the positively charged derivatives are predominate for the highly sensitive detection of carboxylic acids. The APBQ was successfully applied to the selective determination of biological carboxylic acids in human plasma. The bile acids (chenodeoxycholic acid and deoxycholic acid) and several saturated (stearic acid and palmitic acid) and unsaturated free fatty acids (oleic acid and linoleic acid) were reasonably determined by HPLC-MS under the proposed procedure. Based on the results of analyses of human plasma and saliva, the proposed procedure using APBQ seems to be applicable for the qualitative and quantitative analyses of a series of carboxylic acids in biological samples.

Computer-assisted local resection for exostosis osteochondroma of the mandibular condyle.

The purpose of this study was to evaluate the guiding effect of computer-assisted design technique in local resection of exostosis osteochondroma (EOC) from the mandibular condyle. Eight patients diagnosed with EOC through computed tomographic scan were selected from January 2011 to March 2012. SurgiCase CMF 5.0.0.32 software was used for preoperative design. The osteotomy angle, depth, and tumor shape were measured to guide the surgery. The tumors were resected from the stalk 2 mm within the normal tissue to restore the unaffected condyle as much as possible. Postoperative computed tomographic scans showed that the tumors had been resected completely in all patients. The mean (SD) error between the design and the result was 1.82 (1.25) mm. Five patients had more than 1-year follow-up without recurrence. Computer-assisted design is a good way to help local resection of EOC. It can improve the accuracy of tumor resection and keep the unaffected condyle as much as possible for function.

Novel hierarchical carbon microspheres@layered double hydroxides@copper lignosulfonate architecture for polypropylene with enhanced flame retardant and mechanical performances.

Due to the inherent defect of flammability of polypropylene (PP), a novel and highly efficient carbon microspheres@layered double hydroxides@copper lignosulfonate (CMSs@LDHs@CLS) flame retardant was designed and prepared, which was attributed to the strong electrostatic interaction between carbon microspheres (CMSs), layered double hydroxides (LDHs) and lignosulfonate as well as the chelation effect of lignosulfonate on copper ions, and then it was incorporated into the PP matrix. Significantly, CMSs@LDHs@CLS not only observably improved its dispersibility in PP matrix, but also simultaneously achieved excellent flame retardant properties for composites. With the addition of 20.0 % CMSs@LDHs@CLS, the limit oxygen index of CMSs@LDHs@CLS and PP composites (PP/CMSs@LDHs@CLS) reached 29.3 % and achieved the UL-94 V-0 rating. Cone calorimeter tests indicated that the peak heat release rate, total heat release and total smoke production of PP/CMSs@LDHs@CLS composites exhibited declines of 28.8 %, 29.2 % and 11.5 %, respectively, compared with those of PP/CMSs@LDHs composites. These advancements were attributed to the better dispersibility of CMSs@LDHs@CLS in PP matrix and illustrated that CMSs@LDHs@CLS observably reduced fire hazards of PP. The flame retardant property of CMSs@LDHs@CLS might relate to condensed phase flame retardant effect of char layer and catalytic charring of copper oxides.

Iron oxide nanozymes stabilize stannous fluoride for targeted biofilm killing and synergistic oral disease prevention.

Dental caries (tooth decay) is the most prevalent human disease caused by oral biofilms, affecting nearly half of the global population despite increased use of fluoride, the mainstay anticaries (tooth-enamel protective) agent. Recently, an FDA-approved iron oxide nanozyme formulation (ferumoxytol, Fer) has been shown to disrupt caries-causing biofilms with high specificity via catalytic activation of hydrogen peroxide, but it is incapable of interfering with enamel acid demineralization. Here, we find notable synergy when Fer is combined with stannous fluoride (SnF 2 ), markedly inhibiting both biofilm accumulation and enamel damage more effectively than either alone. Unexpectedly, our data show that SnF 2 enhances the catalytic activity of Fer, significantly increasing reactive oxygen species (ROS) generation and antibiofilm activity. We discover that the stability of SnF 2 (unstable in water) is markedly enhanced when mixed with Fer in aqueous solutions without any additives. Further analyses reveal that Sn 2+ is bound by carboxylate groups in the carboxymethyl-dextran coating of Fer, thus stabilizing SnF 2 and boosting the catalytic activity. Notably, Fer in combination with SnF 2 is exceptionally effective in controlling dental caries in vivo , preventing enamel demineralization and cavitation altogether without adverse effects on the host tissues or causing changes in the oral microbiome diversity. The efficacy of SnF 2 is also enhanced when combined with Fer, showing comparable therapeutic effects at four times lower fluoride concentration. Enamel ultrastructure examination shows that fluoride, iron, and tin are detected in the outer layers of the enamel forming a polyion-rich film, indicating co-delivery onto the tooth surface. Overall, our results reveal a unique therapeutic synergism using approved agents that target complementary biological and physicochemical traits, while providing facile SnF 2 stabilization, to prevent a widespread oral disease more effectively with reduced fluoride exposure.

Iron oxide nanozymes stabilize stannous fluoride for targeted biofilm killing and synergistic oral disease prevention.

Dental caries is the most common human disease caused by oral biofilms despite the widespread use of fluoride as the primary anticaries agent. Recently, an FDA-approved iron oxide nanoparticle (ferumoxytol, Fer) has shown to kill and degrade caries-causing biofilms through catalytic activation of hydrogen peroxide. However, Fer cannot interfere with enamel acid demineralization. Here, we show notable synergy when Fer is combined with stannous fluoride (SnF2), markedly inhibiting both biofilm accumulation and enamel damage more effectively than either alone. Unexpectedly, we discover that the stability of SnF2 is enhanced when mixed with Fer in aqueous solutions while increasing catalytic activity of Fer without any additives. Notably, Fer in combination with SnF2 is exceptionally effective in controlling dental caries in vivo, even at four times lower concentrations, without adverse effects on host tissues or oral microbiome. Our results reveal a potent therapeutic synergism using approved agents while providing facile SnF2 stabilization, to prevent a widespread oral disease with reduced fluoride exposure.

Diels-Alder derivatization for sensitive detection and characterization of conjugated linoleic acids using LC/ESI-MS/MS.

The utility of Diels-Alder derivatization with 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) for liquid chromatography/electrospray ionization tandem mass spectrometry of conjugated linoleic acids (CLAs) was examined. PTAD rapidly reacted with the CLAs, and the resulting derivatives were highly responsive in electrospray ionization mass spectrometry operating in the positive-ion mode. The derivatives produced characteristic product ions during tandem mass spectrometry, which enabled the sensitive detection [limit of detection 18 fmol (signal-to-noise ratio of 5)] and the identification of the conjugated diene position. The PTAD derivatization also significantly increased the reversed-phase liquid chromatography separation selectivity for the most biologically active CLA isomers: cis-9,trans-11-CLA and trans-10,cis-12-CLA. The PTAD derivatization was applied to analyses of food and biological samples; the major CLAs in milk and beef fat samples were successfully identified, and trace amounts of CLAs in human saliva were detected with a simple pretreatment and short analysis time.

Simultaneous determination of DL-lactic acid and DL-3-hydroxybutyric acid enantiomers in saliva of diabetes mellitus patients by high-throughput LC-ESI-MS/MS.

A simultaneous determination method for the enantiomers of chiral carboxylic acids by the combination of ultraperformance liquid chromatography and mass spectrometry (UPLC-MS/MS) has been developed. (S)(+)-1-(2-Pyrrolidinylmethyl)-pyrrolidine (S-PMP) was used as the derivatization reagent for the high-throughput determination of biological chiral carboxylic acids, i.e., lactic acid (LA) and 3-hydroxybutyric acid (HA). The S-PMP efficiently reacted with the carboxylic acids under mild conditions at room temperature in the presence of 2,2'-dipyridyl disulfide and triphenylphosphine. The resulting S-PMP derivatives were highly responsive in the electrospray ionization (ESI)-MS operating in the positive-ion mode and gave characteristic product ions during the MS/MS, which enabled the sensitive detection using selected reaction monitoring. The derivatization was effective for the enantiomeric separation of the chiral carboxylic acids, and the resolution values of DL-LA and DL-HA were 4.91 and 9.37, respectively. Furthermore, a rapid separation of the derivatives of DL-LA and DL-HA within 7 min was performed using the UPLC system. The limits of detection on the column were in the low femtogram range (5-12 fg). The proposed procedure was successfully applied for the determination of the D- and L-isomers of LA and HA in the saliva of diabetes mellitus (DM) patients and healthy volunteers. The D-LA in DM patients was clearly higher than that in normal subjects. The derivatization followed by UPLC-ESI-MS/MS enabled the enantiomeric separation and detection of trace amounts of LA and HA in human saliva with a simple pretreatment and small sample volume.