Differential Refractive Index Research Articles

Asymmetric nanofluidic grating detector for differential refractive index measurement and biosensing

Measuring small changes in refractive index can provide both sensitive and contactless information on molecule concentration or process conditions for a wide range of applications. However, refractive index measurements are easily perturbed by non-specific background signals, such as temperature changes or non-specific binding. Here, we present an optofluidic device for measuring refractive index with direct background subtraction within a single measurement. The device is comprised of two interdigitated arrays of nanofluidic channels designed to form an optical grating. Optical path differences between the two sets of channels can be measured directly via an intensity ratio within the diffraction pattern that forms when the grating is illuminated by a collimated laser beam. Our results show that no calibration or biasing is required if the unit cell of the grating is designed with an appropriate built-in asymmetry. In proof-of-concept experiments we attained a noise level equivalent to ∼10-5 refractive index units (30 Hz sampling rate, 4 min measurement interval). Furthermore, we show that the accumulation of biomolecules on the surface of the nanochannels can be measured in real-time. Because of its simplicity and robustness, we expect that this inherently differential measurement concept will find many applications in ultra-low volume analytical systems, biosensors, and portable devices.

Characterization of a water soluble, hyperbranched arabinogalactan from yacon (Smallanthus sonchifolius) roots

Yacon (Smallanthus sonchifolius Poepp. & Endl.) roots are largely grown in Andean countries and have attracted recent interest due to their antioxidant and prebiotic effects. Yacon is typically consumed as a fruit due to its sweet taste and juiciness. The macromolecular properties of an aqueous extract of yacon are investigated using asymmetric flow field-flow fractionation (AF4) coupled to UV, multiangle light scattering (MALS) and differential refractive index (dRI) detection. The method allows for determination of molar mass and size over the size distribution. Three major populations were found of which one strongly dominates in concentration. Through collection of fractions from AF4, carbohydrate composition and glycosidic linkage analysis for the dominating population was performed. The results show that the dominating population consists of a highly branched arabinogalactan (type 2) with a molar mass of approximately 1-2·105g/mol, a hydrodynamic radius of approximately 6-10nm and a relatively high apparent density (approx. 70-150kg/m3).

Analysis of β-glucan molar mass from barley malt and brewer’s spent grain with asymmetric flow field-flow fractionation (AF4) and their association to proteins

β-Glucan benefits are related with its molar mass and it would be of interest to better understand how this parameter can be changed by processing and variety for design of food with specific health effects. For this purpose, extracts from barley malts and brewers' spent grain, processed at different conditions, were analysed regarding β-glucan content, molar mass, and protein content. Molar mass distribution was assessed using asymmetric flow field-flow fractionation (AF4) with multiangle light scattering (MALS), differential refractive index (dRI) and fluorescence (FL) detection. β-Glucan was detected in a wide molar mass range, <2000 to approximately 6.7×106g/mol. Differences in molar masses were more noticeable between barley varieties and steeping malting conditions than by mashing of malt. Barley products processed to preserve β-glucan contained more β-glucan of high molar mass with potential to shift the fermentation site to the distal colon. Enzymatic degradation of proteins indicated presence of aggregates containing β-glucan and protein.

Differential refractive index sensor based on photonic molecules and defect cavities.

We present a novel differential refractive index sensor prototype based on a matrix of photonic molecules (PM) of soda-lime glass cylinders (εc = 4.5) and two defect cavities. The measured and simulated spectra in the microwave range (8-12 GHz) show a wide photonic stop band with two localized states: the reference state, bound to a decagonal ring of cylinders and the sensing state, bound to the defect cavities. The defect mode is very sensitive to the permittivity of the material inserted in the cavity while the state in the PM remains unperturbed. We find that the response of the sensor is linear. These results can be extrapolated to the visible range due to scale invariance of Maxwell equations.

Erratum to: Quantitation and Risk Assessment of Chemical Adulterants in Milk Using UHPLC Coupled to Photodiode Array and Differential Refractive Index Detectors

Milk can be adulterated illegitimately with formaldehyde, melamine, urea, and sugars for preservation, protein content enhancement, and taste improvement, respectively. In the present study, formaldehyde, melamine, urea, and sugars (fructose, glucose, lactose, sucrose) were analyzed using ultra-fast high performance liquid chromatography in 30 fresh and pasteurized milk samples. Satisfactory chromatographic retention and peak shape investigated and method has been validated by means of recovery experiments in samples spiked at two levels. Average recoveries ranged between 95–105 % with excellent precision; RSD lower than 5 % was achieved. Among the tested samples, formaldehyde was detected in all samples while seven and 20 samples were positive for melamine and sucrose, respectively. Monitoring of these adulterants in milk could help to reduce the human health hazards. Exposure to formaldehyde, melamine, and urea from consumption of milk was estimated, and these were less than 3.3, 1, and 700 μg/mL, respectively, which is an oral reference dose suggested by the World Health Organization and US Food and Drug Administration.

Enzymatic cross-linking of carboxymethylpullulan grafted with ferulic acid

Carboxymethylpullulan (CMP) has been modified in a two-step grafting reaction of ferulic acid (FA). Acid adipic dihydrazyde (ADH) was first reacted with FA activated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC). Then the product of this first reaction was reacted with CMP (activated with EDC). Grafted polysaccharides structure was confirmed by FTIR and 1H NMR spectroscopy. Analyses by size-exclusion chromatography (SEC) coupling on-line with a multi-angle light scattering detector (MALS), a viscometer and a differential refractive index detector (DRI) (SEC/MALS/DRI/Visco) showed that associations between FA moieties occurred due to hydrophobic interactions. The grafting rates of FA were determined by the Folin-Ciocalteu method and were found between 1.0% and 11.2% (mol/mol anhydroglucose unit). The CMP-FA were then enzymatically cross-linked with laccase from Pleurotus ostreatus. The crosslinking reactions were followed by rheological measurements, demonstrating the influence of laccase concentration on kinetics. Elastic modulus and swelling rates of hydrogels depends on FA content only for low values.

Quantitation and Risk Assessment of Chemical Adulterants in Milk Using UHPLC Coupled to Photodiode Array and Differential Refractive Index Detectors

Quantitation and Risk Assessment of Chemical Adulterants in Milk Using UHPLC Coupled to Photodiode Array and Differential Refractive Index Detectors

Characterization of Low-Density Polyethylene in Dibutoxymethane by High-Temperature Gel Permeation Chromatography with Triple Detection

The study presents the possibility to replace the 1,2,4-trichlorobenzene (TCB) for the analysis by gel permeation chromatography (GPC) of low-density polyethylenes (LDPE) with dibutoxymethane (DBM, butylal), a halogen free and less toxic solvent. The molecular weight distributions and the viscosity plots were measured for commercial LDPE samples solubilized in TCB, and DBM, using a GPC system with triple detection (light scattering, differential refractive index and viscometer). Similar results were obtained in both solvents for the molecular weight and long chain branching distributions of the analyzed resins, thus confirming the possibility to replace TCB with DBM for the analysis by high-temperature GPC of all types of polyolefins.

A Useful Methodology for Determining the Compaction Degree of Single‐Chain Nanoparticles by Conventional SEC

The folding/collapse of linear single chains to single‐chain nanoparticles (SCNPs) is a field of intense activity. Current synthetic methods allow obtaining from sparse SCNP morphologies in solution – resembling those observed in intrinsically disordered proteins – to compact SCNP conformations, more akin to those displayed by globular enzymes. A useful characterization technique in the field of SCNPs is “conventional” size‐exclusion chromatography (SEC) with differential refractive index (DRI) detection, providing rapid and convincing evidence of SCNP formation through comparison of the SEC traces of the precursor and the SCNPs. Upon SCNP formation, a noticeable shift of the SEC curve toward longer retention times (i.e., lower hydrodynamic sizes) is observed. However, quantitative information embedded in the SEC curve about the actual “compaction degree” of the SCNPs has not yet been extracted to distinguish between sparse and globular SCNPs. Here, a useful methodology for quantifying the compaction degree of SCNPs by SEC with conventional DRI detector is introduced allowing: i) construction of theoretical SEC curves corresponding to the collapse of a linear precursor having a log‐normal molecular weight distribution to either sparse or globular SCNPs and ii) analysis of experimental SEC curves to estimate the actual morphology of synthetic SCNPs in solution.

Theoretical approximations and experimental extinction coefficients of biopharmaceuticals.

UV spectrophotometric measurement is a widely accepted and standardized routine analysis for quantitation of highly purified proteins; however, the reliability of the results strictly depends on the accuracy of the employed extinction coefficients. In this work, an experimental estimation of the differential refractive index (dn/dc), based on dry weight measurements, was performed in order to determine accurate extinction coefficients for four biotherapeutic proteins and one synthetic copolymer after separation in a size-exclusion ultra-performance liquid chromatograph coupled to an ultraviolet, multiangle light scattering and refractive index (SE-UPLC-UV-MALS-RI) multidetection system. The results showed small deviations with respect to theoretical values, calculated from the specific amino acid sequences, for all the studied immunoglobulins. Nevertheless, for proteins like etanercept and glatiramer acetate, several considerations, such as glycan content, partial specific volume, polarizability, and higher order structure, should be considered to properly calculate theoretical extinction coefficient values. Herein, these values were assessed with simple approximations. The precision of the experimentally obtained extinction coefficients, and its convergence towards the theoretical values, makes them useful for characterization and comparability exercises. Also, these values provide insight into the absorbance and scattering properties of the evaluated proteins. Overall, this methodology is capable of providing accurate extinction coefficients useful for development studies.

Characterization of ultrahigh-molecular weight cationic polyacrylamide using frit-inlet asymmetrical flow field-flow fractionation and multi-angle light scattering

In this study, frit inlet asymmetrical flow field-flow fractionation (FlFFF) with multi-angle light scattering (MALS) and differential refractive index (DRI) detection is utilized for size separation, determination of molecular weight (MW), and conformation of ultrahigh-MW (107–109 g/mol) cationic polyacrylamides (C-PAMs), a class of water-soluble copolymers based on acrylamide and vinyl-type comonomers with quaternary ammonium cations that are widely used in wastewater treatment and in paper industries. Linear and branched C-PAM copolymers prepared in two different polymerization methods (solution and emulsion) from varying amounts of crosslinking agent and initiator were size fractionated by FlFFF with field-programming. It was found experimentally that the linear copolymers from both polymerization methods were less than 108 g/mol in MW with compact, nearly spherical structures, while the branched C-PAM copolymers from the emulsion polymerization showed a significant increase in average MW up to ∼109g/mol, which was about 20-fold greater than those from the solution method, and the branched copolymers had more compact or shrunken conformations. While both linear and branched copolymers less than 108 g/mol MW were well resolved in an increasing order of MW (normal mode), it was noted that branched copolymers prepared through emulsion polymerization exhibited significantly larger MWs of 108–109 g/mol and eluted in the steric/hyperlayer mode, in which the elution order is reversed in an extreme run condition (strong initial field strength followed by a fast field decay during programming).

Conventional and Microwave-Assisted Extraction of Mucilage from Opuntia ficus-indica Cladodes: Physico-Chemical and Rheological Properties

Cactus pear cladodes processing has potential value for mainstream industries and is equally important for marginal rural communities in arid regions. This work is focused on physico-chemical and rheological properties of Opuntia ficus-indica (OFI) peeled cladodes extracted by conventional method “CE” and using microwave-assisted extraction “MAE.” MAE gave the highest yield extraction (8.13 %, w/w) within the lowest extraction time (500 W/7 min) and provided more protein (×1.03) and carbohydrates (×1.51) than CE. The monosaccharides detected by gas chromatography were arabinose, galactose, rhamnose, xylose, and galacturonic acid. The dialyzed mucilage solution characterized by SEC/MALS/VD/DRI (size-exclusion chromatography coupled with online multi-angle light scattering, viscometer detectors, and differential refractive index) revealed fractions with molecular weight (M w ) ranging from 15.3–15.7 × 106 g mol−1 for the CE extracts and about 16.7–17.5 × 106 g mol−1 for the MAE extracts. Dynamic oscillatory testing has been used to study the rheological properties of mucilage solution within the concentration of 0.50–3.00 % (w/v) at 25 °C. The rheological profiles of CE and MAE showed similar behavior. At low frequency and low mucilage concentration ( G"). If safely controlled, alternative energies like microwaves could extract soluble polymers with comparable properties to commercial ones, conventionally extracted.

Size-exclusion chromatography of ultrahigh molecular weight methylcellulose ethers and hydroxypropyl methylcellulose ethers for reliable molecular weight distribution characterization

Size-exclusion chromatography (SEC) coupled with multi-angle laser light scattering (MALLS) and differential refractive index (DRI) detectors was employed for determination of the molecular weight distributions (MWD) of methylcellulose ethers (MC) and hydroxypropyl methylcellulose ethers (HPMC) having weight-average molecular weights (Mw) ranging from 20 to more than 1,000kg/mol. In comparison to previous work involving right-angle light scattering (RALS) and a viscometer for MWD characterization of MC and HPMC, MALLS yields more reliable molecular weight for materials having weight-average molecular weights (Mw) exceeding about 300kg/mol. A non-ideal SEC separation was observed for cellulose ethers with Mw>800kg/mol, and was manifested by upward divergence of logM vs. elution volume (EV) at larger elution volume at typical SEC flow rate such as 1.0mL/min. As such, the number-average molecular weight (Mn) determined for the sample was erroneously large and polydispersity (Mw/Mn) was erroneously small. This non-ideality resulting in the late elution of high molecular weight chains could be due to the elongation of polymer chains when experimental conditions yield Deborah numbers (De) exceeding 0.5. Non-idealities were eliminated when sufficiently low flow rates were used. Thus, using carefully selected experimental conditions, SEC coupled with MALLS and DRI can provide reliable MWD characterization of MC and HPMC covering the entire ranges of compositions and molecular weights of commercial interest.

Direct Comparison of IR and DRI Detector for HT‐GPC of Polyolefins

SummaryMolar mass distribution (MMD) of polyolefins is routinely analyzed by gel permeation chromatography (GPC), also known as size exclusion chromatography (SEC), practiced in different methods and detector systems. The filter‐based infrared detector (IR) has become a preferred alternative to the traditional use of differential refractive index detector (DRI) for high temperature GPC (HT‐GPC) when applied to polyolefins. The main reason is that a modern multiple band filter‐based IR detector can be used to determine the distribution of co‐monomer along the molar mass distribution in a copolymer, in a single HT‐GPC analysis directly without the need of additional analytical procedures. In addition to that, the performance of the modern IR detector when used as primary concentration detector in the HT‐GPC instrument, in either the single or the multiple detection schemes, proves to be superior to that of DRI detector. This is due to the high signal‐to‐noise ratio achievable by the modern IR detector and its excellent baseline stability. Such high detection quality can be attributed to the inherent robustness of IR spectroscopy against external factors such as GPC temperature and flow rate variations. The impact of that higher detection performance on the final processed analytical results of polymer MMD and molar mass averages among others, is the focus of this current investigation. Different application scenarios of industrial interest have been selected to show a direct comparison of both the IR and DRI detectors installed on a same GPC instrument.

Determination of the refractive index of glucose-ethanol-water mixtures using spectroscopic refractometry near the critical angle.

A spectroscopic refractometer was used to investigate the dispersion curves of ethanol and D-glucose solutions in water near the critical angle; here, the reflectivity was measured using a white source. Dispersion curves were obtained in the 320-1000 nm wavelength range with a resolution better than 10(-4) for the refractive index, n. The differential refractive index is measured as a function of wavelength, and a simple expression is proposed to obtain the refractive index of the glucose-ethanol-water ternary system. Using this expression, combined with the experimental differential refractive index values, the concentrations of individual components can be calculated.

On-chip integrated differential optical microring refractive index sensing platform based on a laminar flow scheme.

We propose an on-chip integrated differential optical microring refractive index sensing platform which leverages laminar flow conditions. Close spacing between a sensing and a reference resonator, and sharing the same microfluidic channel allows the two resonators to experience similar environmental disturbances, such as temperature fluctuations and fluidic-induced transients, achieving reliable and sensitive sensing performance. We obtain a noise floor of 80.0 MHz (0.3 pm) and a bulk refractive index sensitivity of 17.0 THz per refractive index unit (RIU) (64.2 nm/RIU), achieving a limit of detection of 1.4×10(-5) RIU in a 30 min and an 8°C window.

Characterization of Polypropylene in Dibutoxymethane by High-Temperature Gel Permeation Chromatography with Triple Detection

This study presents the possibility of replacing the 1,2,4-trichlorobenzene (TCB) recommended by ASTM D 6474 for the analysis by gel permeation chromatography (GPC) of polypropylenes with dibutoxymethane (DBM, butylal), a halogen-free and less toxic solvent. The molecular weight distributions as well as the K and alpha parameters of the Mark-Houwink-Sakurada equation were measured for different types of commercial polypropylene samples solubilized in TCB and DBM, using a GPC system with triple detection (light scattering, differential refractive index, and viscometer). For the analyzed resins, covering typical applications of polypropylene, the GPC method in DBM provided results similar to those obtained in TCB.

Miniature temperature sensor with germania-core optical fiber.

A miniature all-fiber temperature sensor is demonstrated by using a Michelson interferometer formed with a short length of Germania-core, silica-cladding optical fiber (Ge-fiber) fusion-spliced to a conventional single-mode fiber (SMF). Thanks to the large differential refractive index of the Ge-fiber sensing element, a reasonably small free spectral range (FSR) of 18.6 nm is achieved even with an as short as 0.9 mm Ge-fiber that may help us increase the measurement accuracy especially in point sensing applications and, at the same time, keep large measurement temperature range without overlapping reading problem. Experimental results show that high sensitivity of 89.0 pm/°C is achieved and the highest measurement temperature is up to 500°C.

Towards determination of absolute molar mass of cellulose polymer by size exclusion chromatography with mulitple angle laser light scattering detection

The study focuses on determination of a set of crucial parameters for molar mass calculation of cellulose from the results of size exclusion chromatography coupled with multiple angle laser light scattering (MALLS) and differential refractive index (DRI) detectors. In the present work, cellulose has been derivatised to obtain cellulose tricarbanilate (CTC) soluble in tetrahydrofuran (THF). The parameters of Rayleigh scattering in the MALLS detector: refractive index increment (dn/dc) and second virial coefficient (A2) of CTC in THF were determined for laser wavelength 658nm. In order to avoid errors resulting from cellulose derivatisation by-products present in the CTC solution, the so called “on-line” method of measuring dn/dc and A2 was applied. Based on the A2 determination, its influence on cellulose molar mass calculations and cellulose molecular dimensions were critically assessed. The latter includes evaluation of artificially aged cellulose towards conceivable branching by conformation plot analysis.

Characterization and Study of Properties in a Polar Solvent of a Functionalized and Quaternized Poly(dimethylsiloxane-co-methyl-hydridosiloxane)

Poly(dimethylsiloxane- co-methyl-hydridosiloxane) (PDMSMHS) with a OSi(CH 3) 2/OSi(HCH 3) molar ratio of 21:1 was functionalized with dimethylallylamine (DMAA) via catalytic hydrosilylation using Karstedt’s catalyst and then quaternized with 1-iodooctane. The functionalized and quaternized co-polysiloxanes (PDMSMHS-1 and PDMSMHS-QAS) were characterized using 1H, 13C and 29Si NMR. The physicochemical properties of the polymeric systems (the differential refractive index increment (dn/dc), the square root of the mean square radius of gyration \(( ^{1/2})\), the average molecular weight (M), and the second virial coefficient (A 2)) were measured in ethanol using gel permeation chromatography/light scattering (GPC/LS). The low percentage (3–4 %) of amino side groups has an unexpected effect on the co-polymers physicochemical properties because the A 2 experimental value for PDMSMHS-1 and PDMSMHS-QAS (8.87×10−3and 1.51×10−2molmLg −2, respectively) indicates that ethanol is a good solvent.