Determination In Milk Research Articles

Physicochemical evaluation of sodium hydroxide solutions for lactic acid determinations in milk

Physicochemical evaluation of sodium hydroxide solutions for lactic acid determinations in milk

Simultaneous reflectance and fluorescence measurements for portable formaldehyde determination in milk using a multi-channel spectrometer sensor

This study introduces a novel optical method for formaldehyde determination in milk, based on the hypothesis that simultaneous reflectance and fluorescence measurements can enhance detection sensitivity compared to traditional methods. We aimed to address the challenge of accurately measuring low concentrations of formaldehyde in milk, a crucial issue for food safety. By employing a multi-channel spectrometer sensor and exploiting the reaction of formaldehyde with acetylacetone to form 3,5-diacetyl-1,4-dihydrolutidine (DDL), we measured reflectance of DDL at 415 nm and fluorescence at 515 nm. The method demonstrated linearity (0.1–4 mg L−1 for reflectance, 0.1–3 mg L−1 for fluorescence) with detection limits of 0.027 mg L−1 (reflectance) and 0.030 mg L−1 (fluorescence). We successfully determined formaldehyde in milk samples (46 to 114 μg L−1) and observed a 60 % reduction in formaldehyde concentrations. This research underscores the importance of heat treatment in ensuring food safety.

DNA-directed immobilization fluorescent immunoarray for multiplexed antibiotic residue determination in milk.

The presence of antibiotic residues in cow's milk entails high risk for consumers, the dairy industry, and the environment. Therefore, the development of highly specific and sensitive screening tools for the rapid and cost-effective identification of traces of these compounds is urgently needed. A multiplexed screening platform utilizing DNA-directed immobilization (DDI) was developed aiming to detect three classes of antibiotic residues (fluoroquinolones, sulfonamides, and tylosin) prevalently found in milk. Throughout this work, each oligonucleotide sequence was conjugated to a different hapten molecule, while the three complementary strands were immobilized in 24 independent microarray chips on a single glass slide. First, the array was incubated with the pool of hapten-oligonucleotide conjugate site encoded the signal through DNA hybridization. Next, commercial milk samples were incubated with the cocktail of monoclonal antibodies following a secondary fluorophore-labeled antibody which was required for fluorescent readout. Direct sample detection was achieved in milk diluting 20 times in assay buffer. The limits of detection (LODs) reached were 1.43µgkg-1, 1.67µgkg-1, and 0.89µgkg-1 for TYLA, STZ, and CIP, respectively, which represented in raw milk 7.15µgkg-1, 8.35µgkg-1, and 4.45µgkg-1 for TYLA, STZ, and CIP, respectively, that are below the EU regulatory limits. Cross-reactivity profiles were evaluated against the family of structurally related antibiotics in order to demonstrate the capability to detect antibiotics from the same family of compounds. A pre-validation study was performed by spiking 20 blind samples above and below the maximum residue limits established by the EU guidelines. The system was successfully implemented towards randomized sample classification as compliant or non-compliant. The proposed DDI-based immunoarray provides a fast and cost-effective alternative to obtain semi-quantitative information about the presence of three veterinary residues simultaneously in milk samples.

A dual-response ratiometric fluorescent sensor for oxytetracycline determination in milk and mutton samples

Owing to the adverse effects of oxytetracycline (OTC) residues on human health, it is of great importance to construct a rapid and effective strategy for OTC detection. Herein, we developed a dual-response fluorescence sensing platform based on molybdenum sulfide quantum dots (MoS2 QDs) and europium ions (Eu3+) for ratiometric detection of OTC. The MoS2 QDs, synthesized through an uncomplicated one-step hydrothermal approach, upon OTC integration into the MoS2 QDs/Eu3+ sensing system, exhibit a significant quenching of blue fluorescence due to the inner filter effect (IFE), simultaneously enhancing the distinct red emission of Eu3+ at 624 nm, a phenomenon attributed to the antenna effect (AE). This sensor demonstrates exceptional selectivity and sensitivity towards OTC, characterized by a linear detection range of 0.2–10 μM and a notably low detection limit of 2.21 nM. Furthermore, we achieved a visual semi-quantitative assessment of OTC through the discernible fluorescence color transition from blue to red under a 365 nm ultraviolet lamp. The practical applicability of this sensor was validated through the successful detection of OTC in milk and mutton samples, underscoring its potential as a robust tool for OTC monitoring in foodstuffs to safeguard food safety.

Electrochemical detection platform based on graphene-zinc oxide composite nanomaterials for 17β-estradiol determination in milk

This research presents the development of an innovative electrochemical detection platform using a graphene-zinc oxide (G-ZnO) nanocomposite for the quantification of 17β-estradiol in milk. The G-ZnO nanocomposite, synthesized via a facile one-pot method, exhibited a unique wrinkled structure with a high specific surface area of 143 m²/g. Characterization through SEM, TEM, and XRD confirmed the effective integration of ultrafine ZnO nanoparticles onto graphene sheets. The electrochemical performance of the G-ZnO modified glassy carbon electrode was evaluated, revealing a notably low detection limit of 8.3 nM for 17β-estradiol, a significant improvement over traditional sensors. The sensor demonstrated a wide linear range of 0.05–100 μM and exceptional reproducibility with less than 3.5 % RSD over repeated measurements. Recovery studies in spiked milk samples yielded highly accurate results, with recovery percentages ranging from 97.8 % to 104.6 %. These findings underscore the potential of G-ZnO nanocomposites in creating sensitive, efficient, and reliable sensors for healthcare and environmental applications.

Protein Determination in Milk: Urgent Issues

Protein Determination in Milk: Urgent Issues

Microplate-based 3D-printed image box for urea determination in milk by digital image colorimetry.

In this study, we describe a rapid and high-throughput smartphone-based digital colorimetric method for determining urea in milk. A compact and cost-effective 3D-printed image box microplate-based system was designed to measure multiple samples simultaneously, using minimal sample and reagent volumes. The apparatus was applied for the quantification of urea in milk based on its reaction with p-dimethylaminobenzaldehyde (DMAB). The predictive performance of calibration was evaluated using RGB and different colour models (CMYK, HSV, and CIELAB), with the average blue (B) values of the RGB selected as the analytical signal for urea quantification. Under optimized conditions, a urea concentration linear range from 50 to 400 mg L-1 was observed, with a limit of detection (LOD) of 15 mg L-1. The values found with the smartphone-based DIC procedure are in good agreement with spectrophotometric (spectrophotometer and microplate treader) and reference method (mid-infrared spectroscopy) values. This proposed approach offers an accessible and efficient solution for digital image colorimetry, with potential applications for various target analytes in milk and other fields requiring high-throughput colorimetric analysis.

Exploiting digital images and videos for urea determination in milk based on enzymatic hydrolysis monitoring

As an endogenous species and a milk adulterant, urea determination is important for quality control in dairy industry. This work proposes a novel, simple, and cost-effective method to determine urea by the analysis of digital images and videos. Urea hydrolysis under urease catalysis releases ammonia and the increase in pH makes feasible the photometric measurements based on the dissociation of the phenol red acid-base indicator. Digital images were acquired by a smartphone camera under controlled conditions and intensities of the reflected radiation were converted to the RGB color system using a free app (PhotoMetrix®). Digital videos were exploited for procedure optimization and evaluation of matrix effects from RGB values obtained with ImageJ® software. A linear response was achieved within 2.0–25.0 mg L-1 urea (R2 =0.996), with a coefficient of variation (n = 10) of 1.6%, and limits of detection (95% confidence level) and quantification of 0.5 and 1.5 mg L-1, respectively. Recoveries from 95% to 112% were estimated from samples spiked with urea and results for commercial milk samples achieved by the standard additions method agreed with the reference values at the 95% confidence level. The procedure is applicable to whole, semi-skim, and skim milk either processed using pasteurization or under ultra-heat treatment.

Fluoride Determination in Milk, Soy, and Water-Based Products Using Ion-Selective Electrode and Direct Measurement Technique: Single-Laboratory Validation, First Action 2022.05.

The AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals issued a call for methods for the determination of fluoride in infant formula. Standard Method Performance Requirement (SMPR®) 2014.016 was approved. A single-laboratory validation (SLV) of a fluoride ion-selective electrode (F-ISE) method was completed. Five powder samples were reconstituted, and hydrochloric acid was added to dissolve and release any bound fluoride. Samples included infant and adult nutritional products made from milk, whey, or soy, containing intact, partially hydrolyzed, or hydrolyzed proteins. Sodium citrate buffer was added to complex any interfering ions and adjust pH and ionic strength. Samples were analyzed in duplicate over 6 days using F-ISE. Results were calculated based on one of two least-squares (log fit) standard curves (0.02 to 0.1 µg/100 g or 0.1 to 2.0 µg/100 g). The LOQ for the method was determined to be 3.3 µg/100 g, which was far below the minimum analytical value of 30 µg/100 g required in the SMPR. The fluoride concentrations ranged from approximately 3 to 14 µg/100 g reconstituted powder and the intermediate precision, RSDr, ranged from 0.7 to 12.4%. Although all of these results are out of scope for the SMPR due to their low fluoride, it should be noted that the RSDr for formulas with fluoride concentrations ranging from approximately 9 to 14 µg/100 g ranged from 0.7 to 4.3%, and the RSDr for the two formulas with fluoride concentrations at or below the theoretical detection limit of the method were only approximately10 and 12.4%. Recoveries ranged from 94 to 98% for samples spiked at three levels with NIST-traceable standard solutions, meeting the SMPR. The F-ISE method meets the SMPR and was adopted as a First Action Official MethodSM. The fluoride ion-selective electrode method presented here is affordable and easy to run. The addition of acid to the samples dissolves any minerals which may bind the fluoride.

Studying the concentration of xenobiotics in milk and developing the biosensor method for their rapid determination

In this article the content of toxic xenobiotics (heavy metals and pesticides) in cow milk collected from 5 districts of Eastern Kazakhstan was examined and their cumulative properties were determined. The content of organochlorine pesticides (HCCH, DDT) was not detected in the analyzed milk. The content of mercury and arsenic in milk samples does not exceed the maximum allowable concentration (0.005 and 0.05 mg/kg, respectively). The content of cadmium above the maximum allowable concentration (0.03 mg/kg) was found in milk sampled from Shemonaikha and Katon-Karagai districts. The content of lead and zinc above the maximum allowable concentration (0.1 and 5.0 mg/kg, respectively) was found in milk samples taken from all 5 studied districts. The content of copper above the maximum allowable concentration (1.0 mg/kg) was found in milk samples collected from 4 districts under study (Borodulikha, Beskaragai, Shemonaikha and Katon-Karagai). Based on the analysis of information data the need to develop an accelerated method of determining toxic xenobiotics in milk was substantiated. The basic directions of modernization of the biosensor for determination of cadmium and lead salts in milk and dairy products were selected. A new approach to the process of immobilization of the enzyme on the surface of a substrate for cadmium and lead salts determination in milk has been developed. The efficiency of using a polymeric plate with a graphite conducting layer as a basis for the enzyme biosensor was established.

LED-Based Desktop Analyzer for Fat Content Determination in Milk.

In dairy, there is a growing request for laboratory analysis of the main nutrients in milk. High throughput of analysis, low cost, and portability are becoming critical factors to provide the necessary level of control in milk collection, processing, and sale. A portable desktop analyzer, including three light-emitting diodes (LEDs) in the visible light region, has been constructed and tested for the determination of fat content in homogenized and raw cow's milk. The method is based on the concentration dependencies of light scattering by milk fat globules at three different wavelengths. Univariate and multivariate models were built and compared. The red channel has shown the best performance in prediction. However, the joint use of all three LED signals led to an improvement in the calibration model. The obtained preliminary results have shown that the developed LED-based technique can be sufficiently accurate for the analysis of milk fat content. The ways of its further development and improvement have been discussed.

Microextraction of Tigecycline Using Deep Eutectic Solvents and Its Determination in Milk by LC-MS/MS Method.

The occurrence of tigecycline (TGC), a new first glycylcycline antibiotic residues in food products harmfully influences potential human consumers health. Therefore, analysts are forced to develop new microextraction methods connected with modern extractants for effective isolation of this compound. For this purpose, deep eutectic solvents (DES) as the extraction media were used. Liquid-liquid microextraction (LLME) of tigecycline from milk samples with application of the hydrophobic deep eutectic solvents: decanoic acid:thymol (1:1), thymol:camphor (2:1), dodecanoic acid:menthol (2:1), and dodecanoic acid:dodecanol (1:1) was developed. The studied samples were subjected to a deproteinization process using trichloroacetic acid solution and acetonitrile. The optimal microextraction parameters, molar ratio of DES components, amount of extraction solvents, pH of milk sample, shaking, and centrifugation time, were chosen. Tigecycline in the obtained microextracts of deep eutectic solvents was analyzed using a liquid chromatographic technique connected with a tandem mass spectrometry (LC-MS/MS) system. The limits of detection and quantification values for TGC determination followed by DES-LLME-LC-MS/MS method were in the 1.8 × 10-11 mol L-1 (0.01 μg kg-1) to 4.0 × 10-9 mol L-1 (2.28 μg kg-1) and 5.5 × 10-11 mol L-1 (0.03 μg kg-1) to 1.2 × 10-8 mol L-1 (6.84 μg kg-1) ranges, respectively. The RSD values of precision were in the range 1.4-7.8% (intraday) and 5.4-11.7% (interday). The developed procedures were used for the determination of tigecycline in different bovine milk samples.

Mach-Zehnder Interferometric Immunosensor for Detection of Aflatoxin M1 in Milk, Chocolate Milk, and Yogurt.

Aflatoxin M1 (AFM1) is detected in the milk of animals after ingestion of aflatoxin B1-contaminated food; since 2002, it has been categorized as a group I carcinogen. In this work, a silicon-based optoelectronic immunosensor for the detection of AFM1 in milk, chocolate milk, and yogurt has been developed. The immunosensor consists of ten Mach-Zehnder silicon nitride waveguide interferometers (MZIs) integrated on the same chip with the respective light sources, and an external spectrophotometer for transmission spectra collection. The sensing arm windows of MZIs are bio-functionalized after chip activation with aminosilane by spotting an AFM1 conjugate with bovine serum albumin. For AFM1 detection, a three-step competitive immunoassay is employed, including the primary reaction with a rabbit polyclonal anti-AFM1 antibody, followed by biotinylated donkey polyclonal anti-rabbit IgG antibody and streptavidin. The assay duration was 15 min with limits of detection of 0.005 ng/mL in both full-fat and chocolate milk, and 0.01 ng/mL in yogurt, which are lower than the maximum allowable concentration of 0.05 ng/mL set by the European Union. The assay is accurate (% recovery values 86.7-115) and repeatable (inter- and intra-assay variation coefficients <8%). The excellent analytical performance of the proposed immunosensor paves the way for accurate on-site AFM1 determination in milk.

PH-responsive dual-enzyme mimics based on hollow metal organic framework-derivatives β-Co(OH)2 for multiple colorimetric assays.

Ahollow metal organic framework derivative β-Co(OH)2has been prepared, which possesses oxidase and peroxidase-like activities. Oxidase-like activity is derived from the generation of free radicals, and peroxidase-like activity is related to the electron transfer process. Unlike other nanozymes with dual enzyme-like activities, β-Co(OH)2 possesses pH-responsive enzyme-like activities, among which the β-Co(OH)2 exhibits superior oxidase and peroxidase-like activities under pH of 4 and 6, respectively, which could avoid mutual interference between multiple enzymes. Based on the phenomenon that enzyme-like activities of β-Co(OH)2 can catalyze colorless TMB to generate blue oxidized TMB (oxTMB) with absorption peak at 652 nm, the sensors integrating total antioxidant capacity and H2O2 quantification were developed. The oxidase-like activity-based colorimetric system has a sensitive response to ascorbic acid, Trolox, and gallic acid, in which the limit of detection for those antioxidant substances was 0.54 μM, 1.26 μM, and 14.34 μM, respectively. The sensors based on peroxidase-like activity hadlow limit of detection of 1.42 μM for H2O2 and alinear range of 5-1000 μM. Theproposed method can be well applied to the detection of the total antioxidant capacity of kiwi, Vc tables, orange and tea extract with high accuracy, and H2O2 determination in milk and glucose detection in beverages with satisfactory recovery (within 97-106%).

A novel approach for lactose determination in cow's milk exploiting smartphone-based digital-image photometry.

Lactose, the main carbohydrate in cow's milk, may cause health problems for consumers with intolerance. Lactose determination in milk is hindered by the matrix complexity and lack of chromophore groups. Chromatography, volumetric, and spectrophotometric approaches involving chemical derivatization are time-consuming and require laborious sample preparation, which is incompatible with the high analytical demand. In this context, a novel approach is presented for lactose determination in milk exploiting smartphone-based digital-image photometry. It was based on a modification of the Benedict's method, involving formation of the violet Cu(I)/2,2'-biquinoline-4,4'-dicarboxylate (BCA) complex instead of the copper(I) oxide precipitate, aiming at improvement of sensitivity and precision. Sample pretreatment and analyte derivatization were performed in Eppendorf tubes with minimal reagent amounts and a smartphone camera was used for image acquisition under controlled conditions. Measurements were based on the RGB color system, taking channel G as the analytical response because of the complementarity with the color of the complex. Under the optimized conditions, the proposed procedure yielded a linear response up to 20 mg L-1 (r = 0.999), with a limit of detection of 1.5 mg L-1, which is compatible with determination of lactose in milk and dairy products categorized with low content of the sugar. The procedure takes less than 10 min, with a coefficient of variation of 3.0% (n = 12) and consumes as low as 160 μg Cu and 430 μg BCA per determination, thus being a more practical, fast, cost-effective, and environmental friendly analytical method.

Development and validation of HPLC-FLD method for aflatoxin M1 determination in milk and dairy products

Abstract Milk and dairy products are the most consumed foods in human diet and their safety is in the attention centre of control authorities. Aflatoxin M1 (AFM1) is a dangerous toxin that can occur in milk and dairy products as a metabolite formed from aflatoxin B1 contained in contaminated animal feed. Therefore, the aim of this study was to develop and validate a reliable method for the determination of AFM1 content in milk and dairy products based on HPLC with fluorescence detection employing immunoaffinity columns (IAC) pre-treatment. Optimal chromatographic separation of AFM1 was achieved using a water/acetonitrile mixture (80/20, v/v) as a mobile phase, column with C18 stationary phase maintained at 25 °C, and fluorescence detection at excitation wavelengths of 360 nm and emission of 440 nm. Efficacy of AFM1 extraction from the samples was found to be influenced by the elution agent composition. The best results were obtained using 1.25 mL of acetonitrile/methanol (3/2, v/v) and 1.25 mL of water. Validation parameters of the proposed method met the criteria set by the European legislation with the limits of detection and quantification at 0.002 and 0.007 µg/kg, respectively. Also, suitability of the method was confirmed by its application for AFM1 determination in certified reference material. Finally, the method was applied for AFM1 determination in 25 milk and dairy products collected in Slovakia; the AFM1 content was below the limit of quantification. It was concluded that the method is suitable for AFM1 content monitoring in milk and dairy products.

Acute phase proteins in cattle and swine: A review

The major acute phase proteins (APPs) in cattle are haptoglobin (Hp) and serum amyloid A (SAA), and in swine, are Hp, SAA, C-reactive protein (CRP), and Pig major acute phase protein (Pig-MAP). Many methodologic assays are presently available to measure these parameters, which are still being improved to increase their specificity, sensitivity, user-friendliness, and economic availability. In cattle, the main applications are the diagnosis and monitoring of frequent diseases such as mastitis and metritis in dairy cows and respiratory problems in young calves. In pigs, APPs are useful in the control of bacterial and viral infections, and they may be used at the slaughterhouse to monitor subclinical pathologies and improve food safety. The utility of APP in animal production must not be forgotten; optimization of protocols to improve performance, welfare, and nutrition may benefit from the use of APPs. Other sample types besides serum or plasma have potential uses; APP determination in milk is a powerful tool in the control of mastitis, saliva is a non-invasive sample type, and meat juice is easily obtained at the slaughterhouse. Increasing our knowledge of reference intervals and the influence of variables such as age, breed, sex, and the season is important. Finally, worldwide harmonization and standardization of analytical procedures will help to expand the use of APPs.

Comparative study of the most commonly used methods for total protein determination in milk of different species and their ultrafiltration products.

Milk ultrafiltration is a widely used membrane filtration process that allows the recuperation of whey proteins in a concentrate high in total solids, which can later be transformed in multiple healthy dairy products with great prospects for the food industry. Protein content is a decisive factor for the technological performance of milk concentrates and currently, the ISO standard method for its determination is Kjeldahl, which is time-consuming and requires specific instrumentation. For this reason, the use of rapid methods to quantify protein would greatly facilitate the monitoring of the milk ultrafiltration process. In this study, the bicinchoninic acid assay (BCA), the detergent compatible Bradford assay and the Dumas method were compared to Kjeldahl protein determination to select a quick and accurate methodology suitable for milk of different species and its ultrafiltration products (retentates and permeates). The protein content obtained from Bradford assay and Dumas method in origin milk and retentate samples was consistent with Kjeldahl values. In contrast, BCA protein levels were significantly different when compared to Kjeldahl and no method was proved to be suitable for protein determination in permeate samples. The use of sodium dodecyl sulfate was also examined to improve protein measurements without success. In comparison with the official method, Bradford assay quantitatively provided the best results, and it would be recommended for a quick, economic and easy determination of total protein content in milk and retentate samples.

Trace determination of Cu(II) in milk samples after preconcentration on Fe3O4@graphene oxide composite based imprinted polymer

AbstractA new tailor‐fitted graphene oxide‐vinyl pyridine ion‐imprinted polymer (mGOVP/IIP), was synthesized using ethylene glycol dimethacrylate as a cross‐linking agent and 2,2′‐azo‐bis‐isobutyronitrile as an initiator to develop dispersive magnetic‐solid‐phase extraction method for the preconcentration of Cu(II) prior to the determination by flame atomic absorption spectrometry. The material was characterized and optimized for various parameters by batch mode, which includes the effect of initial concentration, contact time, temperature, and eluents. Herein, IIP demonstrated a high sorption capacity (63.54 mg g−1) at pH 5.6, with fast kinetics (15 min). The heterogeneity and chemisorption, mechanistic interpretation was explained by the Freundlich isotherm, pseudo‐second‐order kinetic model, respectively. Moreover, the higher selectivity coefficients for Cu(II) in the binary mixturess of concomitant ions signified preferential affinity over similar structural analogues. Additionally, it can be regenerated up to 7 runs and effectively eluted by 1 M HNO3. This developed method exhibited higher accuracy, precision (relative standard deviation &lt;5%), and preconcentration factor (60). The IIP also described potential applications for Cu(II) determination in milk and its by‐products. Overall, the material exhibits ease of access and removal of template ion from the binding sites, cost‐effectiveness, fast mass transfer, promoted competitiveness, reproducibility, reliability, and good regeneration efficiency.

Development of an ultrasensitive molecularly imprinted poly-(ortho-phenylenediamine) based sensor for the determination of melamine adulteration in milk and infant formula.

A sensitive molecularly imprinted poly‐(ortho‐phenylenediamine) electrochemical sensor was fabricated for selective melamine detection in milk and infant formula. The pencil graphite electrode (PGE) was modified by deposition of Au nanoparticles and reduced graphene oxide (RGO) on its surface. The fabrication of the electrode in various stages was monitored using cyclic voltammetry. The immobilized RGO, MIP, and gold nanoparticles on the PGE surface were morphologically characterized by field‐emission scanning electron microscopy (FESEM). Under the optimized conditions, the linear range and the limit of detection (LOD) were 10–17–10–8 M and 2.64 × 10–16 M (S/N = 3), respectively. The prepared sensor exhibited a good reproducibility and repeatability response. The recovery range of melamine‐spiked milk and infant formula was 92.7%–103.9% and 93.5%–105.8%, respectively. The sensor could apply successfully for melamine determination in milk and infant formula samples.