Evaluation of Oil-Absorbing Film for Imprint Desorption Electrospray Ionization Mass Spectrometry Imaging (IDESI-MSI) on Biological Samples.

Imprint Desorption Electrospray Ionization Mass Spectrometry Imaging (IDESI-MSI) has proven to be a robust and reliable tool for chemically imaging biological samples such as fungi, animal tissues, and plants, but the choice of the imprint substrate is crucial. It must effectively transfer maximum amounts of species from the sample while preserving the original spatial distribution of detected molecules. In this study, we explored the potential of utilizing an oil-absorbing film, known for its soft nature and excellent lipophilicity, as an imprint substrate for IDESI-MSI on biological samples. To assess the transfer efficiency of the amounts of molecules and molecular patterns, we conducted experiments using mouse brain tissue. The result shows that more than 90% of the analytes can be transferred to the oil-absorbing film from the original tissue. A comparison of IDESI-MSI results between the oil-absorbing film and the original tissue demonstrates the material's capability to transfer most molecules from the original tissue and retain images of different analytes with high spatial fidelity. We extended our investigation to plant imaging, where we applied IDESI-MSI to a cross-section of okra. The oil-absorbing film exhibited promise in this context as well. These findings suggest that IDESI-MSI utilizing the oil-absorbing film holds potential across various research fields, including biological metabolism, chemistry, and clinical research, making this technique widely applicable.

Comprehensive Characterization of Triterpene Saponins in Rhizoma Panacis Japonici by Offline Two-Dimensional Liquid Chromatography Coupled to Quadrupole Time-of-Flight Mass Spectrometry.

Rhizoma Panacis Japonici (RPJ) is an ancient herbal medicine from China that has long been employed for its medicinal benefits in relieving arthritis physical debility and diverse afflictions. The primary bioactive constituents found in RPJ are triterpene saponins, which exhibit numerous pharmacological actions, including anti-inflammatory, antioxidant, and immunomodulating effects. The present study established a straightforward and effective approach for characterizing triterpene saponins in RPJ. An offline HILIC × RP LC/QTOF-MS method was developed, along with a self-constructed in-house database containing 612 saponins reported in the Panax genus and 228 predicted metabolites. The approach achieved good chromatographic performance in isolating triterpene saponins of RPJ, with the HILIC column as the first dimension (1D) and the BEH C18 column as the second dimension (2D). The developed two-dimensional liquid chromatography system exhibited an orthogonality of 0.61 and a peak capacity of 1249. Detection was performed using a QTOF mass spectrometer in a data-independent manner (MSE) in a negative ion mode. Using the in-house database, the collected MS data were processed by an automatic workflow on UNIFI 1.8.2 software, which included data correction, matching of precursor and product ions, and peak annotation. In this study, 307 saponins were characterized from RPJ and 76 saponins were identified for the first time in Panax japonicus. This research not only enhances our understanding of the chemical characteristics of RPJ but also offers a simple and efficient method for analyzing the complex composition of herbal medicine.

Molecular Mechanism for Converting Carbon Dioxide Surrounding Water Microdroplets Containing 1,2,3-Triazole to Formic Acid.

Spraying water microdroplets containing 1,2,3-triazole (Tz) has been found to effectively convert gas-phase carbon dioxide (CO2), but not predissolved CO2, into formic acid (FA). Herein, we elucidate the reaction mechanism at the molecular level through quantum chemistry calculations and ab initio molecular dynamics (AIMD) simulations. Computations suggest a multistep reaction mechanism that initiates from the adsorption of CO2 by Tz to form a CO2-Tz complex (named reactant complex (RC)). Then, the RC either is reduced by electrons that were generated at the air-liquid interface of the water microdroplet and then undergoes intramolecular proton transfer (PT) or switches the reduction and PT steps to form a [HCO2-(Tz-H)]- complex (named PC-). Subsequently, PC- undergoes reduction and the C-N bond dissociates to generate COOH- and [Tz-H]- (m/z = 69). COOH- easily converts to HCOOH and is captured at m/z = 45 in mass spectroscopy. Notably, the intramolecular PT step can be significantly lowered by the oriented electric field at the interface and a water-bridge mechanism. The mechanism is further confirmed by testing multiple azoles. The AIMD simulations reveal a novel proton transfer mechanism where water serves as a transporter and is shown to play an important role dynamically. Moreover, the transient •COOH captured by the experiment is proposed to be partly formed by the reaction with H•, pointing again to the importance of the air-water interface. This work provides valuable insight into the important mechanistic, kinetic, and dynamic features of converting gas-phase CO2 to valuable products by azoles or amines dissolved in water microdroplets.

Three-dimensional chromatin analysis reveals Sp1 as a mediator to program and reprogram HPV-host epigenetic architecture in cervical cancer.

Human papillomavirus (HPV) is predominantly associated with HPV-related cancers, however, the precise mechanisms underlying the HPV-host epigenetic architectures in HPV carcinogenesis remain elusive. Here, we employed high-throughput chromosome conformation capture (Hi-C) to comprehensively map HPV16/18-host chromatin interactions. Our study identified the transcription factor Sp1 as a pivotal mediator in programming HPV-host interactions. By targeting Sp1, the active histone modifications (H3K27ac, H3K4me1, and H3K4me3) and the HPV-host chromatin interactions are reprogrammed, which leads to the downregulation of oncogenes located near the integration sites in both HPV (E6/E7) and the host genome (KLF5/MYC). Additionally, Sp1 inhibition led to the upregulation of immune checkpoint genes by reprogramming histone modifications in host cells. Notably, humanized patient-derived xenograft (PDX-HuHSC-NSG) models demonstrated that Sp1 inhibition promoted anti-PD-1 immunotherapy via remodeling the tumor immune microenvironment in cervical cancer. Moreover, single-cell transcriptomic analysis validated the enrichment of transcription factor Sp1 in epithelial cells of cervical cancer. In summary, our findings elucidate Sp1 as a key mediator involved in the programming and reprogramming of HPV-host epigenetic architecture. Inhibiting Sp1 with plicamycin may represent a promising therapeutic option for HPV-related carcinoma.

Superfast Formation of C(sp2 )-N, C(sp2 )-P, and C(sp2 )-S Vinylic Bonds in Water Microdroplets.

We report examples of C(sp2 )-N, C(sp2 )-S, and C(sp2 )-P bond-forming transformations in water microdroplets at room temperature and atmospheric pressure using N2 as a nebulizing gas. When an aqueous solution of vinylic acid and amine is electrosprayed (+3 kV), the corresponding C(sp2 )-N product is formed in a single step, which was characterized using mass spectrometry (MS) and tandem mass spectrometry (MS2 ). The scope of this reaction was extended to other amines and other unsaturated acids, including acrylic (CH2 =CHCOOH) and crotonic (CH3 CH=CHCOOH) acids. We also found that thiols and phosphines are viable nucleophiles, and the corresponding C(sp2 )-S and C(sp2 )-P products are observed in positive ion mode using MS and MS2 .

Noninvasive Detection of Skin Cancer by Imprint Desorption Electrospray Ionization Mass Spectrometry Imaging.

We report a technique for the noninvasive detection of skin cancer by imprint desorption electrospray ionization mass spectrometry imaging (DESI-MSI) using a transfer agent that is pressed against the tissue of interest. By noninvasively pressing a tape strip against human skin, metabolites, fatty acids, and lipids on the skin surface are transferred to the tape with little spatial distortion. Running DESI-MSI on the tape strip provides chemical images of the molecules on the skin surface, which are valuable for distinguishing cancer from healthy skin. Chemical components of the tissue imprint on the tape strip and the original basal cell carcinoma (BCC) section from the mass spectra show high consistency. By comparing MS images (about 150-µm resolution) of same molecules from the tape strip and from the BCC section, we confirm that chemical patterns are successfully transferred to the tape stripe. We also used the technique to distinguish cherry angiomas from normal human skin by comparing the molecular patterns from a tape strip. These results demonstrate the potential of the imprint DESI-MSI technique for the noninvasive detection of skin cancers as well as other skin diseases before and during clinical surgery.

The Efficacy of Budesonide, Levocabastine, and Their Combination in Treatment of Vasomotor Rhinitis.

INTRODUCTION: Relevant studies have demonstrated that glucocorticoids and antihistamines, such as budesonide and azelastine, are effective in the treatment of vasomotor rhinitis, with their combined use being more effective than that of a single drug. The aim of this study was to assess the improvement in the symptoms of patients following the combined administration of these drugs. METHODS: We conducted a single-center randomized study on 42 patients. Participants were randomly treated with budesonide, levocabastine hydrochloride, or their combination for 2 weeks. The visual analog scale (VAS) score and levels of eosinophil cationic protein (ECP), histamine (HA), leukotriene B4 (LTB4), and vasoactive intestinal peptide (VIP) in nasal secretions were evaluated before and after treatment. RESULTS: The symptoms of patients were improved in all 3 treatment groups compared with those before treatment. Following combined treatment, the improvement in symptoms of nasal obstruction, runny nose, nasal itching, and sneezing was much greater than those in the groups treated with budesonide or levocabastine hydrochloride alone (p = 0.04, 0.004, 0.005, 0.004, respectively). The decreased levels of these inflammatory mediators were significantly different between the different treatment groups. CONCLUSIONS: Budesonide or levocabastine hydrochloride alone improved the nasal symptoms of patients with vasomotor rhinitis and reduced the levels of ECP, HA, LTB4, and VIP in nasal secretions. However, their combination improved the symptoms of patients more significantly than each drug alone.

One-Step, Catalyst-Free Formation of Phenol from Benzoic Acid Using Water Microdroplets.

Benzoic acid dissolved in water is electrosprayed (-4 kV) by using nitrogen gas at a pressure of 120 psi to form ∼10 µm diameter microdroplets. Analysis with mass spectrometry (MS) and tandem mass spectrometry (MS2) of the resulting microdroplets shows the direct formation of phenol via decarboxylation without any catalyst or added reagents. This process represents an ecofriendly, environmentally benign method for producing phenol and related aromatic alcohols from their corresponding aromatic acids. The mechanism of this transformation was unambiguously characterized using mass spectrometry, radical trapping, and 18O labeling.

Microlensed fiber allows subcellular imaging by laser-based mass spectrometry.

Mass spectrometry imaging (MSI) enables the chemical mapping of molecules and elements in a label-free, high-throughput manner. Because this approach can be accomplished rapidly, it also enables chemical changes to be monitored. Here, we describe a protocol for MSI with subcellular spatial resolution. This is achieved by using a microlensed fiber, which is made by grinding an optical fiber. It is a universal and economic technique that can be adapted to most laser-based mass spectrometry methods. In this protocol, the output of laser radiation from the microlensed fiber causes laser ablation of the sample, and the resulting plume is mass spectrometrically analyzed. The microlensed fiber can be used with matrix-assisted laser desorption ionization, laser desorption ionization, laser ablation electrospray desorption ionization and laser ablation inductively coupled plasma, in each case to achieve submicroscale imaging of single cells and biological tissues. This report provides a detailed introduction of the microlensed fiber design and working principles, sample preparation, microlensed fiber ion source setup and multiple MSI platforms with different kinds of mass spectrometers. A researcher with a little background (such as a trained graduate student) is able to complete all the steps for the experimental setup in ~2 h, including fiber test, laser coupling and ion source modification. The imaging time spent mainly depends on the size of the imaging area. It is suggested that most existing laser-based MSI platforms, especially atmospheric pressure applications, can achieve breakthroughs in spatial resolution by introducing a microlensed fiber module.

The prognosis of patients with small cell carcinoma of the cervix: a retrospective study of the SEER database and a Chinese multicentre registry.

BACKGROUND: Small cell carcinoma of the cervix is a rare but poor prognosis pathological type of cervical cancer, for which advice in clinical guidelines is unspecific. We therefore aimed to investigate the factors and treatment methods that affect the prognosis of patients with small cell carcinoma of the cervix. METHODS: In this retrospective study, we collected data from the Surveillance, Epidemiology, and End Results (SEER) 18 registries cohort and a Chinese multi-institutional registry. The SEER cohort included females diagnosed with small cell carcinoma of the cervix between Jan 1, 2000, and Dec 31, 2018, whereas the Chinese cohort included women diagnosed between Jun 1, 2006, and April 30, 2022. In both cohorts, eligibility was limited to female patients older than 20 years with a confirmed diagnosis of small cell carcinoma of the cervix. Participants who were lost to follow-up or those for whom small cell carcinoma of the cervix was not the primary malignant tumour were excluded from the multi-institutional registry, and those with an unknown surgery status (in addition to those for whom small cell carcinoma of the cervix was not the primary malignant tumour) were excluded from the SEER data. The primary outcome of this study was overall survival (length of time from the date of first diagnosis until the date of death from any cause, or the last follow-up). Kaplan-Meier analysis, propensity score matching, and Cox-regression analyses were used to assess treatment outcomes and risk factors. FINDINGS: 1288 participants were included in the study; 610 in the SEER cohort and 678 in the Chinese cohort. Both univariable and multivariable Cox regression analysis (SEER hazard ratio [HR] 0·65 [95% CI 0·48-0·88], p=0·0058; China HR 0·53 [0·37-0·76], p=0·0005) showed that surgery was associated with a better prognosis. In subgroup analyses, surgery remained a protective factor for patients with locally advanced disease in both cohorts (SEER HR 0·61 [95% CI 0·39-0·94], p=0·024; China HR 0·59 [0·37-0·95]; p=0·029). Furthermore, the protective effect of surgery was observed among patients with locally advanced disease after propensity score matching in the SEER cohort (HR 0·52 [95% CI 0·32-0·84]; p=0·0077). In the China registry, surgery was associated with better outcomes in patients with stage IB3-IIA2 cancer (HR 0·17 [95% CI 0·05-0·50]; p=0·0015). INTERPRETATION: This study provides evidence that surgery improves outcomes of patients with small cell carcinoma of the cervix. Although guidelines recommend non-surgical methods as first-line treatment, patients with locally advanced disease or stage IB3-IIA2 cancer might benefit from surgery. FUNDING: The National Key R&D Program of China and the National Natural Science Foundation of China.

A developmental gradient reveals biosynthetic pathways to eukaryotic toxins in monocot geophytes.

Numerous eukaryotic toxins that accumulate in geophytic plants are valuable in the clinic, yet their biosynthetic pathways have remained elusive. A lead example is the >150 Amaryllidaceae alkaloids (AmAs) including galantamine, an FDA-approved treatment for Alzheimer's disease. We show that while AmAs accumulate to high levels in many tissues in daffodils, biosynthesis is localized to nascent, growing tissue at the base of leaves. A similar trend is found for the production of steroidal alkaloids (e.g. cyclopamine) in corn lily. This model of active biosynthesis enabled elucidation of a complete set of biosynthetic genes for the production of AmAs. Taken together, our work sheds light on the developmental and enzymatic logic of diverse alkaloid biosynthesis in daffodil. More broadly, it suggests a paradigm for biosynthesis regulation in monocot geophytes where plants are protected from herbivory through active charging of newly formed cells with eukaryotic toxins that persist as aboveground tissue develops.

One-Step Formation of Pharmaceuticals Having a Phenylacetic Acid Core Using Water Microdroplets.

The properties of water microdroplets strikingly differ from bulk water. Using room-temperature water microdroplets, we find that toluene can react with CO2 to form phenylacetic acid in one step without any catalyst with negative high voltage applied at the sprayer source. The chemical components of these microdroplets are identified by mass spectrometry, and product structures are confirmed by tandem mass spectrometry. In this manner, we generate three drug molecules in a single step: 4-aminophenylacetic acid (epithelial peptide transporter PepT1 inhibitor), 3,4-dihydroxyphenylacetic acid (dopamine metabolite neurotransmitter), and phenylacetic acid (sodium salt form; treatment of urea cycle disorder). Mechanistic studies show that benzyl radicals formed from hydroxyl radicals at the water microdroplet interface drive these carboxylation reactions. This water microdroplet chemistry is general, allowing activation and subsequent carboxylation of aryl α-C-H groups.

HPV-CCDC106 integration promotes cervical cancer progression by facilitating the high expression of CCDC106 after HPV E6 splicing.

Human papillomavirus (HPV) integration and high expression of HPV oncogenes (E6 and E7) are important mechanisms for HPV carcinogenesis in cervical cancer. However, the relationship between HPV integration and HPV E6 spliced transcripts, as well as the underlying mechanisms of HPV integration in carcinogenesis after HPV E6 splicing remains unclear. We analyzed HPV-coiled-coil domain containing 106 (CCDC106) integration samples to characterize the roles of HPV integration, E6 spliceosome I (E6*I), and high CCDC106 expression in cervical carcinogenesis. We found that E6 was alternatively spliced into the E6*I transcript in HPV-CCDC016 integration samples with low p53 expression, in contrast to the role of E6*I in preventing p53 degradation in cervical cancer cells. In addition, CCDC106 was highly expressed after HPV-CCDC106 integration, and interacted with p53, resulting in p53 degradation and cervical cancer cell progression in vitro and in vivo. Importantly, when E6*I was highly expressed in cervical cancer cells, overexpression of CCDC106 independently degraded p53 and promoted cervical cancer cell progression. In this study, we explored the underlying mechanisms of HPV-CCDC106 integration in HPV carcinogenesis after HPV E6 splicing, which should provide insight into host genome dysregulation in cervical carcinogenesis.

Catalyst-Free Decarboxylative Amination of Carboxylic Acids in Water Microdroplets.

Previous studies have shown that hydroxyl radicals can be formed at the water-gas surface of water microdroplets. We report the use of in situ generated hydroxyl radicals to carry out an organic transformation in one step, namely, the formation of anilines from aryl acids as well as both ammonia and primary/secondary amines via decarboxylation. Benzoic acids and amines are dissolved in water, and the solution is sprayed to form microdroplets whose chemical contents are analyzed mass spectrometrically. All intermediates and products are determined using mass spectrometry (MS) as well as in some cases tandem mass spectrometry (MS2). These results support the following reaction mechanism: NR2OH, formed via reaction of the amine with •OH, reacts with benzoic acid to form an isocyanate via a Lossen rearrangement. Hydrolysis followed by liberation of CO2 then delivers the aniline product. Notably, the scope of this transformation includes a variety of amines and aromatic acids and enables their conversion into aniline and N-substituted anilines, all in a single step. Additionally, this reaction occurs at room temperature and does not require metal catalysts or organic solvents.

The Objective Assessment of dry Nose.

BACKGROUND: Dry nose (DN) is a common symptom in both patients with rhinitis and healthy individuals; however, it is often overlooked. OBJECTIVE: This study aimed to investigate the characteristics of and propose objective diagnostic criteria for DN. METHODS: This study was conducted from December, 2018 to October, 2021. Patients with complaints of a dry nasal cavity and normal controls were recruited consecutively from the allergy-rhinology outpatient clinic of Beijing TongRen Hospital. Questionnaires were completed by each participant during recruitment to record demographic data. DN test strips were used to evaluate the severity of DN. The length of the strip was recorded at 30 s, 1 min, 2 min, 3 min, 4 min, and 5 min, respectively. Nasal secretions were collected on sponges and allergic status was assessed based on serum sage levels. RESULTS: Twenty (13 men and 7 women) patients with DN and 100 (47 men and 53 women) controls were recruited for the study. The participants' ages ranged from 23 to 73 years (mean = 47.7 years). Nine of the 20 DN patients were diagnosed with vasomotor rhinitis. The weight of the sponges of DN patients was significantly lower than that of controls. At the last time point (5 min), the strips in the control group were significantly longer than those in the DN group. The reference range of 30 s, 1 min, 2 min, 3 min, 4 min, and 5 min of controls was 3.0 mm, 6.0 mm, 10.9 mm, 13.2 mm, 16.8 mm, and 17.0 mm, respectively. CONCLUSIONS: Our study indicated that the strip length less than 17.0 mm at 5 min is a valuable reference for the diagnostic of DN in Beijing.

Direct C(sp3)-N Bond Formation between Toluene and Amine in Water Microdroplets.

Unlike the inertness of bulk water, water microdroplets exhibit some remarkable reactivities. We report that water microdroplets can directly produce stable C7H7+ cations (a combination of benzylic and tropylium cations) from toluene and other substrates at room temperature with a positive voltage (+4 kV) applied to the droplet spray source. The C7H7+ cation and the benzyl radical (C6H5CH2·) are both generated via hydroxyl radicals at the water-gas interface of the microdroplets. The C7H7+ signal is observed directly by mass spectrometry. Dissolved amines (primary, secondary, and tertiary) in the microdroplets can react with both C7H7+ and C6H5CH2· to form the corresponding alkyl C(sp3)-N coupling products in one step, which cannot be achieved in bulk water or other solvents. The products were identified using tandem mass spectrometry (MS2) and 1H NMR spectroscopy. Notably, the direct C(sp3)-N bond formation products were obtained in the absence of a catalyst. In the presence of a radical scavenger, the mass spectra of the C(sp3)-N coupling products are strongly suppressed, which supports the hypothesis that this reaction is driven by hydroxyl radicals generated in the water microdroplets. Taken together, these results show that water microdroplets provide a new method for direct one-step C(sp3)-N bond formation without the need for a metal catalyst.

Spraying Water Microdroplets Containing 1,2,3-Triazole Converts Carbon Dioxide into Formic Acid.

We report the use of 1,2,3-triazole (Tz)-containing water microdroplets for gas-phase carbon dioxide (CO2) reduction at room temperature. Using a coaxial sonic spraying setup, the CO2 can be efficiently captured by Tz and converted to formic acid (HCOOH; FA) at the gas-liquid interface (GLI). A mass spectrometer operated in negative ion mode monitors the capture of CO2 to form the bicarbonate anion (HCO3-) and conversion to form the formate anion (HCOO-). Varied FA species were successfully identified by MS/MS experiments including the formate monomer ([FA - H]-, m/z 45), the dimer ([2FA - H]-, m/z 91; [2FA + Na - 2H]-, m/z 113), the trimer ([3FA - H]-, m/z 137), and some other adducts (such as [FA - H + H2CO3]-, m/z 107; [2FA + Na - 2H + Tz]-, m/z 182). The reaction conditions were systematically optimized to make the maximum conversion yield reach over 80% with an FA concentration of approximately 71 ± 3.1 µM. The mechanism for the reaction is speculated to be that Tz donates the proton and the hydroxide (OH-) at the GLI, resulting in a stepwise yield of electrons to reduce gas-phase CO2 to FA.

Laser Ablation Electrospray Ionization Achieves 5 µm Resolution Using a Microlensed Fiber.

A pulsed (10 Hz) infrared (IR) (1064 nm) laser is focused on a sample surface by means of a microlensed fiber. Analytes desorbed from the surface are captured by charged microdroplets before entering a mass spectrometer. By translating the sample surface, a chemical map is generated with a resolution of 5 µm, defined as the change from 20 to 80% of the analyte signal intensity. As a demonstration of the power of this new imaging technique, analytes from a parsnip root section are imaged and compared with that obtained from conventional laser ablation electrospray ionization mass spectrometry. The improvement in spatial resolution is about a factor of 20.

CAP-YOLO: Channel Attention Based Pruning YOLO for Coal Mine Real-Time Intelligent Monitoring.

Real-time coal mine intelligent monitoring for pedestrian identifying and positioning is an important means to ensure safety in production. Traditional object detection models based on neural networks require significant computational and storage resources, which results in difficulty of deploying models on edge devices for real-time intelligent monitoring. To address the above problems, CAP-YOLO (Channel Attention based Pruning YOLO) and AEPSM (adaptive image enhancement parameter selection module) are proposed in this paper to achieve real-time intelligent analysis for coal mine surveillance videos. Firstly, DCAM (Deep Channel Attention Module) is proposed to evaluate the importance level of channels in YOLOv3. Secondly, the filters corresponding to the low importance channels are pruned to generate CAP-YOLO, which recovers the accuracy through fine-tuning. Finally, considering the lighting environments are varied in different coal mine fields, AEPSM is proposed to select parameters for CLAHE (Contrast Limited Adaptive Histogram Equalization) under different fields. Experiment results show that the weight size of CAP-YOLO is 8.3× smaller than YOLOv3, but only 7% lower than mAP, and the inference speed of CAP-YOLO is three times faster than that of YOLOv3. On NVIDIA Jetson TX2, CAP-YOLO realizes 31 FPS inference speed.

Capture of Hydroxyl Radicals by Hydronium Cations in Water Microdroplets.

Despite the high stability of bulk water, water microdroplets possess strikingly different properties, such as the presence of hydroxyl radicals (OH⋅) at the air-water interface. Previous studies exhibited the recombination of OH⋅ into H2 O2 molecules and the capture of OH⋅ by oxidizing other molecules. By spraying pure water microdroplets into a mass spectrometer, we detected OH⋅ in the form of (H4 O2 )+ that is essentially OH⋅-H3 O+ , a hydroxyl radical combined with a hydronium cation through hydrogen bonding. We also successfully captured it with two OH⋅ scavengers, caffeine and melatonin, and key oxidation radical intermediates that bear important mechanistic information were seen. It is suggested that some previous reactions involving (H4 O2 )+ should be attributed to reactions with OH⋅-H3 O+ rather than with the water dimer cation (H2 O-OH2 )+ .