[Determination of 11 nutrients in liquid milk by ultra-performance liquid chromatography-tandem mass spectrometry].

OBJECTIVE: To establish an ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) method for simultaneous determination of 11 nutritional components(thiamine, riboflavin, nicotinamide, nicotinic acid, pantothenic acid, pyridoxine, pyridoxal, pyridoxamine, biotin, choline, L-carnitine) in liquid milk. METHODS: Milk samples were shaken with 20 mmol/L ammonium formate solution and heated in a water bath at 100 ℃ for 30 min, then incubated with papain and acid phosphatase at 45 ℃ for 16 h, the lower liquid was collected after centrifugation for analysis. UPLC separation was performed on an ACQUITY~(TM) HSS T3(3.0 mm×150 mm, 1.8 µm) column, 2 mmol/L ammonium formate(containing 0.1% formic acid) solution and acetonitrile(containing 0.1% formic acid) were used as mobile phase. Quantitative detection was performed by internal standard method. RESULTS: 11 nutritional components can be effectively separated and detected in 12 min, and the linear correlation coefficients(R~2) were all above 0.995. The limits of detection(LODs) were between 0.05 and 0.50 µg/L, and the limits of quantification(LOQs) were between 0.20 and 1.25 µg/L. The recovery rates of three-level addition were 85.6%-119.3%, and the precision RSDs were between 3.68% and 7.82%(n=6). Based on the detection of 60 liquid milk samples from 5 different animals, it was found that the contents of 11 nutrients in liquid milk from different milk sources were significantly different, but pyridoxine could not be detected. CONCLUSION: The method can quantitatively detect 11 water-soluble nutrients, including free and bound forms, by effective enzymolysis. It is sensitive, reproducible and can meet the needs of quantitative detection.

Impurity-profiling UPLC methods for quantitative analysis of some antiemetics formulated with pyridoxine.

Cyclizine hydrochloride (CYC) and meclozine hydrochloride (MEC) are antihistaminic drugs generally co-formulated with pyridoxine hydrochloride (PYR) to treat nausea and vomiting in pregnancy. Several analytical techniques have been applied for the determination of CYC or MEC with PYR, but determination of CYC impurity; benzhydrol (BEH) or MEC impurity; or 4-chlorobenzophenone (BEP) has not been paid attention to. Therefore, micellar UPLC method is introduced for analysis of ternary mixtures containing PYR together with both CYC and BEH (mixture I) or MEC and BEP (mixture II). Chromatographic separation was achieved using a Hypersil gold C8 column (50 × 2.1 mm, 1.9 µm) using 0.01 M sodium dodecyl sulfate modified to pH 3.5 using phosphoric acid:acetonitrile (45:55 by volume) for mixture I and 0.1% sodium dodecyl sulfate, 0.1% sodium bicarbonate adjusted to pH 2.6 by phosphoric acid:acetonitrile (47:53 by volume) for mixture II as mobile phases. The separated peaks were detected at 230 and 245 nm for mixtures I and II, respectively. The adopted methods were validated in conformance with the International Conference on Harmonization (ICH) recommendations and were properly applied in commercial pharmaceutical formulation analysis. Comprehensive ecological comparison was achieved, confirming a higher ecological value of the presented methods compared to the earlier reported methods.

Nutritional and Functional Evaluation of Inula crithmoides and Mesembryanthemum nodiflorum Grown in Different Salinities for Human Consumption.

The nutritional composition and productivity of halophytes is strongly related to the biotic/abiotic stress to which these extremophile salt tolerant plants are subjected during their cultivation cycle. In this study, two commercial halophyte species (Inula crithmoides and Mesembryanthemum nodiflorum) were cultivated at six levels of salinity using a soilless cultivation system. In this way, it was possible to understand the response mechanisms of these halophytes to salt stress. The relative productivity decreased from the salinities of 110 and 200 mmol L-1 upwards for I. crithmoides and M. nodiflorum, respectively. Nonetheless, the nutritional profile for human consumption remained balanced. In general, I. crithmoides vitamin (B1 and B6) contents were significantly higher than those of M. nodiflorum. For both species, ß-carotene and lutein were induced by salinity, possibly as a response to oxidative stress. Phenolic compounds were more abundant in plants cultivated at lower salinities, while the antioxidant activity increased as a response to salt stress. Sensory characteristics were evaluated by a panel of culinary chefs showing a preference for plants grown at the salt concentration of 350 mmol L-1. In summary, salinity stress was effective in boosting important nutritional components in these species, and the soilless system promotes the sustainable and safe production of halophyte plants for human consumption.

Effect of different drying methods on product quality, bioactive and toxic components of Ginkgo biloba L. seed.

BACKGROUND: Ginkgo biloba seeds are used as a functional food across Asia. However, the presence of toxic compounds has limited their application. In this study, freeze drying, infrared drying, hot-air drying and pulsed-vacuum drying were used to dry G. biloba seeds. A comprehensive analysis was performed on their product quality, antioxidant activities, bioactive and toxic components. RESULTS: Results showed that the drying methods had a significant influence on product quality with freeze drying being superior due to the minimal microstructural damage, followed by infrared drying and pulsed-vacuum drying. Infrared-dried product possessed the strongest antioxidant activities and higher bioactive compound content than hot-air-dried and pulsed-vacuum-dried product. Toxic compounds in fresh G. biloba seeds (ginkgotoxin, ginkgolic acid and cyanide) were reduced markedly by drying. Ginkgotoxin was reduced fourfold, and the contents of acrylamide, ginkgolic acid and cyanide in dried G. biloba seeds were reduced to the scope of safety. Amongst the four drying methods, infrared drying had the shortest drying time, and its product showed higher quality and bioactive compound content, and stronger antioxidant activities. CONCLUSIONS: These findings will offer salient information for selecting a drying method during the processing of ginkgo seeds. Infrared drying could be considered as a multiple-effect drying method in the processing of ginkgo seeds. © 2020 Society of Chemical Industry.

Submerged fermentation of Ginkgo biloba seed powder using Eurotium cristatum for the development of ginkgo seeds fermented products.

BACKGROUND: Ginkgo biloba seeds are well known for the significant curative effects on relieving cough and asthma. However, the development of products from ginkgo seeds still falls behind at present, resulting in a great waste of ginkgo seeds' resource. In this work, submerged fermentation of ginkgo seed powder using Eurotium cristatum was studied to investigate its feasibility as a new processing method. RESULTS: To promote the growth of E. cristatum, the optimum fermentation medium was 80.0 g L-1 of ginkgo seed powder with addition of 5.0 g L-1 calcium chloride (CaCl2 ), 4.0 g L-1 magnesium sulfate (MgSO4 ), 1.25 g L-1 zinc sulfate (ZnSO4 ) and 0.65 g L-1 iron(II) sulfate (FeSO4 ). The optimum fermentation conditions were pH 5.8 ± 0.1, inoculum size 5.1 × 106 CFU mL-1 , liquid medium volume 100 mL in 250-mL Erlenmeyer flask and fermentation 4 days. Through fermentation, the production of lovastatin in fermentation broth could reach up to 32.97 ± 0.17 µg mL-1 and the total antioxidant capacity was improved by more than two-fold. In addition, 40.15% of the ginkgotoxin in ginkgo seed powder was degraded while the entire degradation of ginkgolic acids was obtained. Moreover, fermented ginkgo seed powder suspension presented pleasant fragrances, and the activities of amylase and protease were enhanced to 11.30 ± 0.10 U mL-1 and 23.01 ± 0.20 U mL-1 , respectively. CONCLUSIONS: Submerged fermentation using E. cristatum could significantly enhance the functional value and safety of ginkgo seed powder, and had great potential to become a novel processing method for the development of ginkgo seeds fermented products. © 2020 Society of Chemical Industry.

Relationship between Vitamin Deficiencies and Co-Occurring Symptoms in Autism Spectrum Disorder.

Recently, connections have been made between feeding and eating problems and autism spectrum disorder (ASD) and between autism pathophysiology and diet issues. These could explain some of the mechanisms which have not yet been discovered or are not sufficiently characterized. Moreover, there is an increased awareness for micronutrients in ASD due to the presence of gastrointestinal (GI) problems that can be related to feeding issues. For example, levels of vitamins B1, B6, B12, A and D are often reported to be low in ASD children. Thus, in the present mini review we focused on describing the impact of some vitamins deficiencies and their relevance in ASD patients.

The Use of a Biostimulant Composition to Increase Turkey Viability during the Key Critical Periods of Development.

The paper reports that the treatment of hatching turkey eggs with a mixture composed of colamine, succinic acid, serine, and pyridoxine hydrochloride increased the viability of embryos and reduced incubation wastes. This effect allowed increasing the hatching of turkey poults by 6.73% and the hatchability of eggs, by 4.43%. At the same time, a statistically significant decrease in the key lipid peroxidation products in one-day-old turkey poults was observed. In particular, the content of isolated double bonds decreased 1.47-fold (p < 0.01); diene conjugates, 1.67-fold (p < 0.01); triene conjugates, 1.46-fold (p < 0.05); oxidiene conjugates, 1.48-fold (p < 0.01); and Schiff bases, 1.3-fold compared to the control. All the above-mentioned positively affected survivability in the experimental group, which appeared to be increased by 1% compared to the control.

A short-term stimulation of ethanol enhances the effect of magnetite on anaerobic digestion.

Conductive iron oxides (CIO) have been proved recently to facilitate the anaerobic microbial syntrophy based on the direct interspecies electron transfer (DIET) in batch experiments. However, the effect of CIO was always insignificant in anaerobic digestion (AD) reactor especially when the DIET-based syntrophic partners were absent. In this study, the effect of magnetite on performance of AD system with sucrose as a sole carbon source was investigated, but limited enhancement was achieved during the first 36-day operation. The short-term effect of ethanol addition was further studied in the magnetite-amended AD reactor, and results showed that the AD reactor with 10gFe/L micro-sized magnetite (R3) achieved higher performance of COD removal and methane proportion compared with the other reactors (R1 without magnetite; R2 with 2gFe/L micro-sized magnetite; R4 with 2gFe/L nano-sized magnetite). Meanwhile, the pyridoxine in extracellular polymeric substances (EPS) and conductivity of anaerobic sludge from R3 increased more significantly than those of the others. Analysis of high-throughput sequencing indicated that the abundance of archaea increased in sludge from R3 and Methanosarcina responsible for DIET was dominant (63.64%). Additionally, the abundance of potential electroactive bacteria Chloroflexi in R3 was 7.57-fold, 3.61-fold and 7.37-fold as that of R1, R2 and R4, respectively. These results demonstrated that the electroactive microbes and methanogens could be enriched efficiently in anaerobic sludge via synergetic effect of magnetite addition and ethanol short-term stimulation.

Hydrogen sulphide production by Saccharomyces cerevisiae UCD 522 in a synthetic grape juice medium deficient of thiamin (vitamin B1 ) and/or pyridoxine (vitamin B6 ).

The impacts of thiamin and pyridoxine along with YAN on alcoholic fermentation and hydrogen sulphide production by Saccharomyces cerevisiae were studied. Using a synthetic grape juice medium, three fermentation trials were conducted; (i) 2 × 3 factorial design with thiamin (0, 0·2, or 0·5 mg l-1 ) and YAN (60 or 250 mg l-1 ) as variables, (ii) 2 × 3 factorial design with pyridoxine (0, 0·25, or 0·5 mg l-1 ) and YAN (60 or 250 mg l-1 ) as variables, and (iii) 3 × 3 factorial design with thiamin (0, 0·2 or 0·5 mg l-1 ) and pyridoxine (0, 0·25 or 0·5 mg l-1 ) as variables in media containing 60 mg l-1 YAN. Although the progress of fermentations was affected by thiamin or pyridoxine, YAN had a larger impact than either vitamin. H2 S production was significantly lower with increasing amounts of thiamin in those fermentations under low YAN (60 mg l-1 ) while even lower amounts (<30 µg l-1 ) were produced under high YAN (250 mg l-1 ) with or without the vitamin. The highest amounts of H2 S were synthesized in those fermentations without any pyridoxine (>110 µg l-1 ), with the lowest production in media with pyridoxine and high YAN (<20 µg l-1 ). SIGNIFICANCE AND IMPACT OF THE STUDY: Concentrations of thiamin, pyridoxine and yeast assimilable nitrogen (YAN) influenced the synthesis of hydrogen sulphide (H2 S) by Saccharomyces cerevisiae in a synthetic grape juice medium. With a few exceptions, an increase in the concentration of either vitamin or YAN resulted in less H2 S released. This is the first report to demonstrate that both thiamin and pyridoxine along with YAN affected H2 S production, emphasizing the need to assess yeast nutrients to lower risks of off-odours during fermentation.

Analysis of the Overlapped Electrochemical Signals of Hydrochlorothiazide and Pyridoxine on the Ethylenediamine-Modified Glassy Carbon Electrode by Use of Chemometrics Methods.

In this work, the electrochemical behavior of hydrochlorothiazide and pyridoxine on the ethylenediamine-modified glassy carbon electrode were investigated by differential pulse voltammetry. In pH 3.4 Britton-Robinson (B-R) buffer solution, both hydrochlorothiazide and pyridoxine had a pair of sensitive irreversible oxidation peaks, that overlapped in the 1.10 V to 1.20 V potential range. Under the optimum experimental conditions, the peak current was linearly related to hydrochlorothiazide and pyridoxine in the concentration range of 0.10-2.0 µg/mL and 0.02-0.40 µg/mL, respectively. Chemometrics methods, including classical least squares (CLS), principal component regression (PCR) and partial least squares (PLS), were introduced to resolve the overlapped signals and determine the two components in mixtures, which avoided the troublesome steps of separation and purification. Finally, the simultaneous determination of the two components in commercial pharmaceuticals was performed with satisfactory results.

Simultaneous determination of niacin and pyridoxine at trace levels by using diode array high-performance liquid chromatography and liquid chromatography with quadrupole time-of-flight tandem mass spectrometry.

A highly sensitive and simple diode-array high-performance liquid chromatography and liquid chromatography with quadrupole time-of-flight tandem mass spectrometry method was developed for the simultaneous determination of niacin and pyridoxine in pharmaceutical drugs, tap water, and wastewater samples. To determine the in vivo behavior of niacin and pyridoxine, analytes were subjected to simulated gastric conditions. The calibration plots of the diode-array high-performance liquid chromatography and liquid chromatography with quadrupole time-of-flight tandem mass spectrometry method showed good linearity over a wide concentration range with close to 1.0 correlation coefficients for both analytes. The limit of detection/limit of quantitation values for liquid chromatography quadrupole time-of-flight tandem mass spectrometry analysis were 1.98/6.59 and 1.3/4.4 µg/L for niacin and pyridoxine, respectively, while limit of detection/limit of quantitation values for niacin and pyridoxine in high-performance liquid chromatography analysis were 3.7/12.3 and 5.7/18.9 µg/L, respectively. Recovery studies were also performed to show the applicability of the developed methods, and percentage recovery values were found to be 90-105% in tap water and 94-97% in wastewater for both analytes. The method was also successfully applied for the qualitative and quantitative determination of niacin and pyridoxine in drug samples.

Modified Eu-doped Y2 O3 nanoparticles as turn-off luminescent probes for the sensitive detection of pyridoxine.

Europium-doped yttrium oxide nanoparticles (Y2 O3 :Eu NPs) modified by captopril were prepared in aqueous solution. In this study, we report the effect of pyridoxine hydrochloride on the photoluminescence intensity of Y2 O3 :Eu NPs in pH 7.2 buffer solution. By increasing the pyridoxine concentration, the luminescence intensity of Y2 O3 :Eu NPs is quenched. The results show that this method demonstrates high sensitivity for pyridoxine determination. A linear relationship is observed between 0.0 and 62.0 µM with a correlation coefficient of 0.995 and a detection limit of 0.023 µM.

Quantification of a botanical negative marker without an identical standard: ginkgotoxin in Ginkgo biloba.

A new strategy for the analysis of natural products uses a combination of quantitative (1)H NMR (qHNMR) and adsorbent-free countercurrent separation (CS) methodology to establish a quantification method for ginkgotoxin (4'-O-methylpyridoxine) in Ginkgo biloba preparations. The target analyte was concentrated in a one-step CS process using the ChMWat +2 solvent system (CHCl3-MeOH-H2O, 10:5:5) and subsequently assayed by qHNMR. While commercial G. biloba seeds contained 59 µg of ginkgotoxin per seed, the compound was below the limit of detection (9 ppm) in a typical leaf extract. Due to the enrichment potential and loss-free operation of CS, the combination of CS and qHNMR is a generally suitable approach for threshold assays aimed at quantifying target compounds such as botanical negative markers at the low ppm level. As the proof of principle is demonstrated for relatively small CS capacities (20 mL, 1:40 loading) and modest NMR sensitivity (n = 16, 400 MHz, 5 mm RT probe), the approach can be adapted to quantification at the ppb level. The procedure enables the quantification of a botanical negative marker in the absence of identical reference material, which otherwise is a prerequisite for LC-based assays.

Electrochemical sensing of vitamin B6 (pyridoxine) by adapted carbon paste electrode.

The recent investigation targets to use adapted carbon paste (CP) with copper nanoparticles (CuNs) operating in a phosphate buffer (PBS) medium with a pH range of 5.0-8.0, to synthesize a novel, susceptible, and simple electrochemical sensor for the detection of one of the most important drugs, vitamin B6. Copper (Cu) is one of the most three common essential trace elements found in the bodies of both humans and animals, along with iron and zinc for all crucial physiological and biochemical functions. Its properties, which are assessed using a variety of methods including scanning electron microscopy (SEM), cyclic voltammetry (CV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS), have also drawn a lot of attention recently. We considered the effects of pH, buffer, scan rate, interference, and calibration curve. The susceptible electrode's linear calibration curve encompassed concentration values between 8.88 and 1000.0 µM. The calculated limits of detection and quantification were 32.12 and 107.0 µM, respectively. Furthermore, this method was established in real human urine samples and drug validation which have been shown satisfactory results for vitamin B6 detection.

Comparative ANNs with different input layers and GA-PLS study for simultaneous spectrofluorimetric determination of melatonin and pyridoxine HCl in the presence of melatonin's main impurity.

Melatonin (MLT) has many health implications, therefore it is important to develop specific analytical methods for the determination of MLT in the presence of its main impurity, N-{2-[1-({3-[2-(acetylamino)ethyl]-5-methoxy-1H-indol-2-yl}methyl)-5-methoxy-1H-indol-3-yl]ethyl}acetamide (DMLT) and pyridoxine HCl (PNH) as a co-formulated drug. This work describes simple, sensitive, and reliable four multivariate calibration methods, namely artificial neural network preceded by genetic algorithm (GA-ANN), principal component analysis (PCA-ANN) and wavelet transform procedures (WT-ANN) as well as partial least squares preceded by genetic algorithm (GA-PLS) for the spectrofluorimetric determination of MLT and PNH in the presence of DMLT. Analytical performance of the proposed methods was statistically validated with respect to linearity, accuracy, precision and specificity. The proposed methods were successfully applied for the assay of MLT in laboratory prepared mixtures containing up to 15% of DMLT and in commercial MLT tablets with recoveries of no less than 99.00%. No interference was observed from common pharmaceutical additives and the results compared favorably with those obtained by a reference method.

Vitamin B6: deficiency diseases and methods of analysis.

Vitamin B6 (pyridoxine) is closely associated with the functions of the nervous, immune and endocrine systems. It also participates in the metabolic processes of proteins, lipids and carbohydrates. Pyridoxine deficiency may result in neurological disorders including convulsions and epileptic encephalopathy and may lead to infant abnormalities. The Intravenous administration of pyridoxine to patients results in a dramatic cessation of seizures. A number of analytical methods were developed for the determination of pyridoxine in different dosage forms, food materials and biological fluids. These include UV spectrometric, spectrofluorimetric, mass spectrometric, thin-layer and high-performance liquid chromatographic, electrophoretic, electrochemical and enzymatic methods. Most of these methods are capable of determining pyridoxine in the presence of other vitamins and complex systems in µg quantities. The development and applications of these methods in pharmaceutical and clinical analysis mostly during the last decade have been reviewed.

The importance, prevalence and determination of vitamins B6 and B12 in food matrices: A review.

Vitamins are a vast group of fundamental organic compounds, which are not produced by the human body but are essential for the living organisms' good health. Vitamins B6 and B12 belong to the same group of hydrophilic vitamins. Structurally unrelated, they share the same purpose as essential components for normal cellular operation, growth and development. Vitamin B6 is an enzymatic co-factor that is vital for countless biochemical reactions, and is also important in sugar and fatty acid metabolization. It encompasses three natural and inter-convertible pyridine-derivatives: pyridoxine, pyridoxal and pyridoxamine. Vitamin B12 is a cobalt organometallic complex also indispensable in numerous human physiological functions. It has four bioactive forms: cyanocobalamin, methylcobalamin, hydroxocobalamin and 5'-deoxyadenosylcobalamin, and only a few prokaryotes have the ability to biosynthesize cobalamin. This work reviews the significant aspects of vitamins B6 and B12: their vital roles, consequences of deficit; food sources; and methods of determination and respective matrices, with heavy emphasis on chromatographic techniques developed within the last two decades.

Content and stability of B complex vitamins in commercial cosmetic products.

BACKGROUND: Individual B vitamins have many favorable effects on the skin and are common cosmetic ingredients. However, their formulation is demanding due to stability issues, which consequently affect the products' quality. AIMS: We aimed to determine the quality (labeling accuracy, content determination, and content-related quality control) and stability under long-term and accelerated storage conditions of a representative sample of commercial cosmetics containing the most common B vitamins - nicotinamide, dexpanthenol, pyridoxine, and cyanocobalamin. METHODS: Cyanocobalamin was determined by a previously published stability-indicating HPLC- diode array detector (DAD) method for the simultaneous determination of all hydrophilic vitamins. This method was additionally simplified and adjusted for the time-effective analysis of nicotinamide, dexpanthenol, and pyridoxine. Both methods were properly validated. RESULTS: All labeled B vitamins were present in the 36 tested products, mostly in contents, reported effective on the skin. Thus, a straightforward correlation between vitamin contents and product prices were not observed. The content-related quality control of eight products, which quantitively specify their content, revealed significantly lower nicotinamide contents (47% and 57%) in two products and appropriate or higher nicotinamide (102%-112%) and dexpanthenol (100%-104%) contents than declared in the remaining products. The 6-month long-term and accelerated stability studies demonstrated the products' physical stability, but also revealed dexpanthenol, pyridoxine, and cyanocobalamin degradation, while nicotinamide was mostly stable in the tested products. CONCLUSIONS: The obtained results provide an inside into the quality of commercial vitamin B cosmetics and highlight the importance of stability testing in the formulation of quality, efficient, and safe cosmetics.

Thermal decomposition of pyridoxine: an evolved gas analysis-ion attachment mass spectrometry study.

RATIONALE: Pyridoxine is an important vitamer in food and pharmaceutical products. Heat treatments applied during preparation or storage of the products cause the decomposition of pyridoxine. Identification and understanding of the degradation products of pyridoxine and studying its decomposition kinetics are essential in the preparation and preservation of pyridoxine-containing foods and pharmaceuticals. METHODS: Real-time, non-isothermal decomposition of pyridoxine was studied using evolved gas analysis-Li(+) ion attachment mass spectrometry (EGA-Li(+) IAMS). Arrhenius parameters for the thermal decomposition of pyridoxine were obtained via the total ion monitoring (TIM) curve. RESULTS: Most of the pyridoxine evaporated in molecular form, but the formation of pyridoxal and o-quinone methide, both biologically important species, was also observed from the solid-phase degradation of pyridoxine. The observation of o-quinone methide, a species possessing anticancer activity, was particularly noteworthy due to its chemical instability. The activation energy (E(a) ) for pyridoxine decomposition determined by EGA-IAMS was found to be 20.0 kcal mol(-1) , and the pre-exponential factor (A) was 5.7 × 10(9) min(-1) . CONCLUSIONS: The calculated kinetic parameters are important for predicting the thermal stability of pyridoxine vitamer. The estimated lifetime (t(90%,25°C) ) of 1.7 × 10(-2) years in nitrogen was also obtained from the EGA-IAMS experiment.

Fractional-continuous wavelet transforms and ultra-performance liquid chromatography for the multicomponent analysis of a ternary mixture containing thiamine, pyridoxine, and lidocaine in ampules.

New chemometric approaches based on the application of partial least squares (PLS) and principal component regression (PCR) algorithms with fractional wavelet transform (FWT) and continuous wavelet transform (CWT) are proposed for the spectrophotometric multicomponent determination of thiamine hydrochloride (B1), pyridoxine hydrochloride (B6), and lidocaine hydrochloride (LID) in ampules without any separation step. In this study PLS and PCR techniques were applied to the raw spectral data, FWT-coefficients, and FWT-CWT-coefficients. These calibration models were labeled as Raw-PLS and Raw-PCR, FWT-PLS and FWT-PCR, and FWT-CWT-PLS and FWT-CWT-PCR, respectively. A new ultra-performance liquid chromatographic (UPLC) method was developed for the comparison of the results obtained by applying the chemometric calibration methods. Chromatographic separation and determination of B1, B6, and LID in ampules were performed on an Acquity UPLC BEH C18 column (50x2.1 mm id, 1.7 pm particle size) using gradient elution with a mobile phase consisting of methanol and 0.01 M HCI at a constant flow rate of 0.6 mL/min. These combined chemometric calibrations and UPLC were validated by analyzing various ternary mixtures, B1, B6, and LID. The proposed chemometric approaches (signal processing-multivariate calibrations) and UPLC method were applied to the quantitative multicomponent analysis of marketed ampules containing the vitamins B1 and B6 with LID.