Mortality in septic patients treated with vitamin C: a systematic meta-analysis.

BACKGROUND: Supplementation of vitamin C in septic patients remains controversial despite eight large clinical trials published only in 2020. We aimed to evaluate the evidence on potential effects of vitamin C treatment on mortality in adult septic patients. METHODS: Data search included PubMed, Web of Science, and the Cochrane Library. A meta-analysis of eligible peer-reviewed studies was performed in accordance with the PRISMA statement. Only studies with valid classifications of sepsis and intravenous vitamin C treatment (alone or combined with hydrocortisone/thiamine) were included. RESULTS: A total of 17 studies including 3133 patients fulfilled the predefined criteria and were analyzed. Pooled analysis indicated no mortality reduction in patients treated with vitamin C when compared to reference (risk difference - 0.05 [95% CI - 0.11 to - 0.01]; p = 0.08; p for Cochran Q = 0.002; I2 = 56%). Notably, subgroup analyses revealed an improved survival, if vitamin C treatment was applied for 3-4 days (risk difference, - 0.10 [95% CI - 0.19 to - 0.02]; p = 0.02) when compared to patients treated for 1-2 or > 5 days. Also, timing of the pooled mortality assessment indicated a reduction concerning short-term mortality (< 30 days; risk difference, - 0.08 [95% CI - 0.15 to - 0.01]; p = 0.02; p for Cochran Q = 0.02; I2 = 63%). Presence of statistical heterogeneity was noted with no sign of significant publication bias. CONCLUSION: Although vitamin C administration did not reduce pooled mortality, patients may profit if vitamin C is administered over 3 to 4 days. Consequently, further research is needed to identify patient subgroups that might benefit from intravenous supplementation of vitamin C.

Vitamin C measurement in critical illness: challenges, methodologies and quality improvements.

Background There is renewed interest in high-dose vitamin C interventions in clinical medicine due to its antioxidant properties, safe use and cost-effectiveness. Yet, randomised control trials (RCTs) employing these interventions are failing to include robust analytical methodology and proper sample handling and processing techniques. Consequently, comparisons between studies becomes impossible as there is no metrological traceability and results may be prone to pre-analytical errors. Content Through published vitamin C stability studies, method comparison papers and data from vitamin C external quality assurance programs, an assessment was made on the functionality of current methods for critically ill patient samples. Summary Data was obtained from two external quality assurance programs, two papers assessing sample stability and interlaboratory agreement and a publication on vitamin C method comparisons. A shift from spectrophotometric and enzymatic methodologies to high performance liquid chromatography (HPLC) greatly improved the variability and interlaboratory agreement. Therefore, the current analytical performance of vitamin C HPLC methodologies are acceptable for the requirements of a high-dose vitamin C RCTs. Outlook Recommendations across the total testing process of vitamin C have been provided to improve the quality of the results. The harmonisation of sample handling and processing procedures will further improve the reliability of current analytical methodologies.

Zwitterionic-hydrophilic interaction liquid chromatography for l-ascorbic acid 2-phosphate magnesium, a raw material in cell therapy.

l-ascorbic acid 2-phosphate magnesium (APMg) salt is a vitamin C derivative frequently used in cell culture media for research purposes. It is also used as a raw material in the GMP-manufacturing of gene-, cell- and tissue advanced therapy medicinal products (ATMPs). However, quality methods are currently lacking. Therefore, a LC method was developed, based on hydrophilic interaction (HILIC)-ion exchange (IE) mixed-mode liquid chromatography. The final method consisted of an isocratic system with 15 mM KH2PO4 buffer (pH 2.5 with HCl) acetonitrile (30:70, v/v) mobile phase on a zwitterionic HILIC column, containing an hydrophilic ligand embedded cation-exchange functionality and a surface anion-exchange group. A flow rate of 0.4 mL/min and UV detection at 240 nm was applied. The assay method of APMg was validated, obtaining adequate linearity (R2 = 0.999), precision (RSD of 0.49%) and accuracy (overall recovery of 100.4%). The developed method was successfully applied on five currently marketed products from different suppliers, showing different related substance impurity profiles. Using atomic absorption spectroscopy (AAS), magnesium was found to be bound on the stationary phase, requiring a strong mobile phase to rinse the column. Finally, related impurities were identified using MS/MS and high resolution MS, and found to be ascorbic acid as well as ethyl derivatives, which was further confirmed by NMR.

Are the Recommended Dietary Allowances for Vitamins Appropriate for Elderly People?

BACKGROUND: An adequate vitamin intake is essential for a good nutritional status, especially in older women, who are more sensitive to nutritional deficiencies. The American, European and Italian Recommended Dietary Allowances (RDAs) derive mainly from studies on adults, and it is not clear whether they also apply to elderly people. Comparing the RDAs with the actual vitamin intake of a group of healthy older women could help to clarify the real needs of elderly people. OBJECTIVE: Our aim was to compare the American, European, and Italian RDAs with the actual vitamin intake of a group of healthy older women. DESIGN: This was a cross-sectional study. PARTICIPANTS: The study included 286 healthy women aged older than 65 years. MAIN OUTCOME MEASURES: For each micronutrient, the 50th percentile of the distribution of its intake was considered as the average requirement, and the corresponding calculated RDA for our sample was the average requirement×1.2, as recommended by the US Food and Nutrition Board. This calculated RDA was then compared with the American, European, and Italian RDAs. STATISTICAL ANALYSES PERFORMED: Student's t test or the Mann-Whitney test (after checking the normal distribution of the micronutrient) for continuous variables; the χ(2) test for categorical variables. RESULTS: The calculated RDA were 2,230 µg retinol equivalents for vitamin A, 2.8 µg for vitamin B-12, 0.9 mg for thiamin, 1.4 mg for riboflavin, 3.6 mg for pantothenic acid, 1.4 mg for vitamin B-6, 320 µg for folic acid, and 115 mg for vitamin C. CONCLUSIONS: Our findings suggest that the current RDAs are adequate for older women's intake of riboflavin, vitamin B-6, and folic acid, but should be raised for vitamin B-12 and for vitamin C.

Characterization of NIST food-matrix Standard Reference Materials for their vitamin C content.

The vitamin C concentrations in three food-matrix Standard Reference Materials (SRMs) from the National Institute of Standards and Technology (NIST) have been determined by liquid chromatography (LC) with absorbance detection. These materials (SRM 1549a Whole Milk Powder, SRM 1849a Infant/Adult Nutritional Formula, and SRM 3233 Fortified Breakfast Cereal) have been characterized to support analytical measurements made by food processors that are required to provide information about their products' vitamin C content on the labels of products distributed in the United States. The SRMs are primarily intended for use in validating analytical methods for the determination of selected vitamins, elements, fatty acids, and other nutrients in these materials and in similar matrixes. They can also be used for quality assurance in the characterization of test samples or in-house control materials, and for establishing measurement traceability. Within-day precision of the LC method used to measure vitamin C in the food-matrix SRMs characterized in this study ranged from 2.7% to 6.5%.

Thermal stability of vitamin C: thermogravimetric analysis and use of total ion monitoring chromatograms.

The thermal decomposition kinetics and shelf life of vitamin C in nitrogen or air were studied by using thermogravimetric analysis (TGA) and evolved-gas analysis-lithium-ion attachment mass spectrometry (EGA-Li⁺IAMS). Arrhenius parameters obtained via TGA were reported for thermal decomposition. For vitamin C in a nitrogen atmosphere, the activation energy (E(a)) was 25.1 kcal/mol and the pre-exponential factor (A) was 2.5 × 10¹¹ min⁻¹. The kinetic parameters estimated via TGA agreed with values estimated from a pyrogram when the weight loss observed by TGA was shown to be due to gas evolution as a result of decomposition of the compound. Thermal stability was expressed by calculating the time for 10% of the vitamin C to decompose at 25 °C (t(90%,25 °C)). The t(90%,25 °C) for vitamin C obtained via TGA or EGA-Li⁺IAMS was higher in nitrogen (2.0 and 2.0 years, respectively) than in air (1.3 and 1.6 years, respectively). This indicates that the type of atmosphere influences vitamin C stability.

A novel automated electrochemical ascorbic acid assay in the 24-well microtiter plate format.

Automatic ascorbic acid (AA) voltammetry was established in 24-well microtiter plates. The assay used a movable assembly of a pencil rod working, an Ag/AgCl reference and a Pt counter electrode with differential pulse voltammetry (DPV) for concentration-dependent current generation. A computer was in command of electrode (z) and microtiter plate (x, y) positioning and timed potentiostat operation. Synchronization of these actions supported sequential approach of all wells and subsequent execution of electrode treatment procedures or AA voltammetry at defined intervals in a measuring cycle. DPV in well solutions offered a linear current/concentration range between 0.1 and 8.0 mM, a sensitivity of about 1 µA mM(-1) AA, and a detection limit of 50 µM. When used with a calibration curve or standard addition, automated voltammetry of samples with added known amounts of AA demonstrated good recovery rates. Also, the assay achieved the accurate determination of the AA content of vitamin C tablets, a fruit juice and an herbal tea extract. Robotic AA voltammetry has the advantage of conveniently handling multiple samples in a single measuring run without the continuous attention of laboratory personnel. It is a good option when the goal is cost-effective AA screening of sample libraries and has potential for applications in health care and the food processing, cosmetic and pharmaceutical industries.

Wavelet analysis used for spectral background removal in the determination of glucose from near-infrared single-beam spectra.

Wavelet analysis is developed as a preprocessing tool for use in removing background information from near-infrared (near-IR) single-beam spectra before the construction of multivariate calibration models. Three data sets collected with three different near-IR spectrometers are investigated that involve the determination of physiological levels of glucose (1-30 mM) in a simulated biological matrix containing alanine, ascorbate, lactate, triacetin, and urea in phosphate buffer. A factorial design is employed to optimize the specific wavelet function used and the level of decomposition applied, in addition to the spectral range and number of latent variables associated with a partial least-squares calibration model. The prediction performance of the computed models is studied with separate data acquired after the collection of the calibration spectra. This evaluation includes one data set collected over a period of more than 6 months. Preprocessing with wavelet analysis is also compared to the calculation of second-derivative spectra. Over the three data sets evaluated, wavelet analysis is observed to produce better-performing calibration models, with improvements in concentration predictions on the order of 30% being realized relative to models based on either second-derivative spectra or spectra preprocessed with simple additive and multiplicative scaling correction. This methodology allows the construction of stable calibrations directly with single-beam spectra, thereby eliminating the need for the collection of a separate background or reference spectrum.

Hydrophilic interaction liquid chromatography method for the determination of ascorbic acid.

Hydrophilic interaction liquid chromatography (HILIC) method using internal standard for the determination and stability study of ascorbic acid was developed. HILIC method was very fast and simple using the following analytical conditions: ZIC HILIC (150 x 2.1 mm, 3.5 microm) chromatographic column and mobile phase composed of ACN and 50 mM ammonium acetate buffer pH 6.8 (78:22 v/v). Diode array detection was performed and chromatograms were processed at 268 nm, the maximum wavelength of absorbance of ascorbic acid. An extensive stability study of ascorbic acid as a function of various factors including temperature, stabilizing agents, oxygen presence and its concentration in solution was performed in order to gain information about the quantitative influence of individual stability factors. Low temperature and stabilizing agents (o-phosphoric acid and oxalic acid) were found to be key factors enabling substantial enhancement of the stability of ascorbic acid.

High-throughput analysis of vitamin C in human plasma with the use of HPLC with monolithic column and UV-detection.

Vitamin C plays a central role in the body. One of its important functions is its role as an antioxidant, and accurate measurements are important for interpretations of this role. However, its reactive nature and instability complicates the assessment, especially in biological samples. A high-throughput chromatographic method using monolithic column and UV-detection was developed for the assessment of plasma ascorbic acid and total ascorbic acid. The method showed excellent analytical sensitivity, specificity, precision, recovery and linearity during the validation study. The method was used for the assessment of ascorbic acid and total ascorbic acid during several clinical studies.

Screening chemical composition and in vitro antioxidant and antimicrobial activities of the essential oils from Origanum syriacum L. growing in Turkey.

In the present study, essential oil from the leaves of Syrian oreganum [Origanum syriacum L. (Lauraceae)] grown in Turkish state forests of the Dortyol district, Turkey, was obtained by steam distillation. The chemical composition of oil was analysed by GC and GC-MS, and was found to contain 49.02% monoterpenes, 36.60% oxygenated monoterpenes and 12.59% sesquiterpenes. The major components are as follows: gamma-terpinene, carvacrol, p-cymene and beta-caryophyllene. Subsequently, the reducing power, antioxidant and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging activities of the essential oil were studied. The reducing power was compared with ascorbic acid, and the other activities were compared with 2,6-di-tert-butyl-4-methyl phenol (BHT, butylated hydroxytoluene). The results showed that the activities were concentration dependent. The antioxidant activities of the oil were slightly lower than those of ascorbic acid or BHT, so the oil can be considered an effective natural antioxidant. Antimicrobial activities of the essential oil from the leaves of Origanum syriacum was also determined on 16 microorganisms tested using the agar-disc diffusion method, and showed antimicrobial activity against 13 of these.

Use of redox potential modification by gas improves microbial quality, color retention, and ascorbic acid stability of pasteurized orange juice.

The aim of this paper was to study the effect of both redox potential (Eh) and pasteurization of orange juice on stability of color and ascorbic acid, and growth recovery of microorganisms during storage at 15 degrees C for 7 weeks. Three conditions of Eh, +360 mV (ungassed), +240 mV (gassed with N2), and -180 mV (gassed with N2-H2) were applied to orange juice. Both thermal destruction and recovery of sublethally heat-injured cells of Lactobacillus plantarum and Saccharomyces cerevisiae were investigated. While oxidizing conditions were the most effective for thermal destruction of L. plantarum and S. cerevisiae, reducing conditions decreased recovery of heated cells of S. cerevisiae. In addition, gassing the juice with N2 or N2-H2 increased color retention and ascorbic acid stability. The present study demonstrated that juice must be reduced just after the heat treatment in order, firstly, to maximize microbial destruction during pasteurization, and secondly, to prevent the development of microorganisms and stabilize color and ascorbic acid during storage.

Certification of standard reference material 970, ascorbic acid in serum, and analysis of associated interlaboratory bias in the measurement process.

BACKGROUND: The accurate and reproducible measurement of ascorbic acid is essential in delineating the role of ascorbic acid as a diagnostic tool for human disease and for the comparison of data acquired by different laboratories. A stabilized pair of standards of ascorbic acid in human serum, which is compatible with most analytical methods, have been prepared. METHODS: The certification was based on the gravimetric addition of ascorbic acid to metaphosphoric acid-stabilized, ascorbic acid-depleted serum and NIST liquid chromatography-electrochemical measurements. The NIST results were analyzed statistically for homogeneity, and the expanded uncertainty of each SRM was calculated using all of the NIST data. An interlaboratory comparison exercise was also performed. RESULTS: These materials, Standard Reference Material (SRM) 970 Ascorbic Acid in Serum, Level I and Level II, are homogeneous and are certified to contain (10.07 +/- 0.21) and (30.57 +/- 0.28) mmol ascorbic acid/L of solution (expanded uncertainty), respectively. In the interlaboratory comparison (n = 17), the relative SDs for the two materials were 22% and 19%. CONCLUSIONS: Two lots of serum, each containing different amounts of ascorbic acid stabilized in metaphosphoric acid, have been prepared and characterized. Many laboratories provide inaccurate results.

Measurement of ascorbic acid in single human neutrophils by capillary zone electrophoresis with electrochemical detection.

A method has been developed for the determination of ascorbic acid (AA) in individual human neutrophils by capillary zone electrophoresis with electrochemical detection at a carbon fiber microdisk bundle electrode. The natively easily oxidized substances such as glutathione, dopa, dopamine, serotonin, epinephrine do not interfere with the determination of ascorbic acid. A procedure of treating capillaries, which can overcome the influence of the adsorption of the substances in cells on the inner surface wall of the capillary on the migration time and the number of theoretical plates of interests, has been described. The average amount of AA in an individual neutrophil is 0.557 fmol, which is consistent with the literature value.

Estimates of the theoretical maximum daily intake of erythorbic acid, gallates, butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) in Italy: a stepwise approach.

The three recent EU directives which fixed maximum permitted levels (MPL) for food additives for all member states also include the general obligation to establish national systems for monitoring the intake of these substances in order to evaluate their use safety. In this work, we considered additives with primary antioxidant technological function for which an acceptable daily intake (ADI) was established by the Scientific Committee for Food (SCF): gallates, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and erythorbic acid. The potential intake of these additives in Italy was estimated by means of a hierarchical approach using, step by step, more refined methods. The likelihood of the current ADI to be exceeded was very low for erythorbic acid, BHA and gallates. On the other hand, the theoretical maximum daily intake (TMDI) of BHT was above the current ADI. The three food categories found to be main potential sources of BHT were "pastry, cake and biscuits", "chewing gums" and "vegetables oils and margarine"; they overall contributed 74% of the TMDI. Actual use of BHT in these food categories is discussed, together with other aspects such as losses of this substance in the technological process and percentage of ingestion in the case of chewing gums.

Antioxidants: what role do they play in physical activity and health?

Exercise appears to increase reactive oxygen species, which can result in damage to cells. Exercise results in increased amounts of malondialdehyde in blood and pentane in breath; both serve as indirect indicators of lipid peroxidation. However, not all studies report increases; these equivocal results may be due to the large intersubject variability in response or the nonspecificity of the assays. Some studies have reported that supplementation with vitamins C and E, other antioxidants, or antioxidant mixtures can reduce symptoms or indicators of oxidative stress as a result of exercise. However, these supplements appear to have no beneficial effect on performance. Exercise training seems to reduce the oxidative stress of exercise, such that trained athletes show less evidence of lipid peroxidation for a given bout of exercise and an enhanced defense system in relation to untrained subjects. Whether the body's natural antioxidant defense system is sufficient to counteract the increase in reactive oxygen species with exercise or whether additional exogenous supplements are needed is not known, although trained athletes who received antioxidant supplements show evidence of reduced oxidative stress. Until research fully substantiates that the long-term use of antioxidants is safe and effective, the prudent recommendation for physically active individuals is to ingest a diet rich in antioxidants.

Vitamin E, vitamin C, and exercise.

Exercise increases the generation of oxygen free radicals and lipid peroxidation. Strenuous exercise in a person who is unconditioned or unaccustomed to exercise will induce oxidative damage and result in muscle injury. However, aerobic exercise training strengthens the antioxidant defense system by increasing superoxide dismutase. Vitamin C and, especially, vitamin E are shown to decrease the exercise-induced increase in the rate of lipid peroxidation. No ergogenic effects of either vitamin C or E have been shown. Vitamin E was shown to significantly increase circulating neutrophils in older, but not younger, subjects performing eccentric exercise that causes an increase in skeletal muscle damage. In addition to its effect in augmenting the neutrophil response to eccentric exercise, vitamin E causes a greater increase in circulating creatine kinase activity, perhaps indicating increased skeletal muscle repair. Increased vitamin E intake has been associated with enhanced glucose tolerance and insulin action as well as improved lipoprotein status. Future research should examine the combined effects of exercise training and vitamins E and C on these health-related outcomes.

Nonenzymatic elimination of ascorbic acid in clinical samples.

OBJECTIVES: Ascorbic acid interferes significantly in the oxidative reaction of chromogenic reagents by peroxidase and hydrogen peroxide. Currently, ascorbate oxidase is commonly utilized for eliminating the interference of ascorbic acid in the oxidative colorimetric reaction. This enzyme, however, displays several disadvantages, such as high cost, variation from lot to lot, and low stability. We applied a series of commercially available and stable radicals (ascorbic acid quenchers [AAQs]) for nonenzymatic quenching of ascorbic acid in the uricase-based uric acid determination in serum and urine. DESIGN AND METHODS: In order to evaluate the quenching activity of AAQs, a commercially available uric acid detection kit was used. TBA-80FR.NEO biochemical analyzer was utilized for the assay. RESULTS: 4-Hydroxy-2,2,6,6-tetramethyl-1-piperidinyloxy free radical (AAQ-2) was the most effective ascorbic acid quencher among the four stable radicals, and the uric acid assay suffered no interference by AAQ-2. The ascorbic acid quenching ability of 2 mmol/L of AAQ-2 in reagent solution (reagent-I) was > or = 2 U/ml ascorbate oxidase in reagent solution. CONCLUSIONS: AAQ-2 was proven to be a suitable quencher of ascorbic acid in clinical samples.