Three-phase partitioning as an elegant and versatile platform applied to nonchromatographic bioseparation processes.

In recent years, a big trend has been the development of rapid, green, efficient, economical, and scalable approaches for the separation and purification of bioactive molecules from natural sources, which can be used in food, cosmetics, and medicine. As a new nonchromatographic bioseparation technology, three-phase partitioning (TPP) is attracting the attention of a growing number of scientists and engineers. Although a number of studies have been published in the last 40 years regarding the extraction, separation, and purification of numerous bioactive molecules using TPP systems, a background review on TPP partitioning fundamentals and its applications is much needed. Therefore, the present review focuses in detail on the TPP separation process, including the definition of TPP, partitioning mechanisms, parameters for establishing the suitable condition to form precipitate such as concentration of ammonium sulfate, content of tert-butanol, pH and temperature, and the application for separation and purification of protein, enzyme, plant oil, polysaccharide, and other small molecule organic compounds. In addition, the possible directions of future developments in TPP technology are discussed. The review presents a good opportunity, as well as a challenge for scientists, to understand the detailed partitioning rule and to take better use of TPP for the production and separation of various bioactive molecules, which have been intensively applied in the food and medical fields.

Biosynthesized gold nanoparticles that activate Toll-like receptors and elicit localized light-converting hyperthermia for pleiotropic tumor immunoregulation.

Manipulating the tumor immune contexture towards a more active state can result in better therapeutic outcomes. Here we describe an easily accessible bacterial biomineralization-generated immunomodulator, which we name Ausome (Au + [exo]some). Ausome comprises a gold nanoparticle core covered by bacterial components; the former affords an inducible hyperthermia effect, while the latter mobilizes diverse immune responses. Multiple pattern recognition receptors actively participate in Ausome-initiated immune responses, which lead to the release of a broad spectrum of pro-inflammatory cytokines and the activation of effector immune cells. Upon laser irradiation, tumor-accumulated Ausome elicits a hyperthermic response, which improves tissue blood perfusion and contributes to enhanced infiltration of immunostimulatory modules, including cytokines and effector lymphocytes. This immune-modulating strategy mediated by Ausome ultimately brings about a comprehensive immune reaction and selectively amplifies the effects of local antitumor immunity, enhancing the efficacy of well-established chemo- or immuno-therapies in preclinical cancer models in female mice.

Effects of structural and conformational characteristics of citrus pectin on its functional properties.

Ultrasonic degradation has become a fascinating strategy for preparing modified pectin, contributing to the improvement of pectin's functional characteristics. In this study, the impacts of structural and conformational characteristics of original and ultrasound-treated citrus pectins on their functional properties were examined. The results showed that compared with ultrasound-treated pectins, untreated pectin presented higher rheological and gelling properties primarily attributed to its larger weight-average molecular weight (Mw), degree of methoxylation, amount of neutral sugar side chains, and z-average radius of gyration, as well as the more extended flexible-chain conformation. However, the ultrasound-treated pectins had better emulsifying properties than untreated pectin in an oil-in-water emulsion system. These properties, visually suggested by morphology analysis, including enhanced emulsifying capacity, emulsifying stability, reduced mean droplet size and negatively charged zeta potential. Moreover, the Mw and chain conformation of untreated and ultrasound-treated pectins played more decisive roles in their functional properties than the others.

Ultrasonic treatment at different pH values affects the macromolecular, structural, and rheological characteristics of citrus pectin.

Ultrasonic degradation has become a promising strategy for producing modified pectin (MP). In this study, the impact of ultrasonic treatment at various pH values (4.0, 7.0, and 10.0) on the macromolecular, structural and rheological characteristics of citrus pectin was investigated. Results demonstrated that ultrasonic irradiation at the higher pH led to larger reductions in the intrinsic viscosity and weight-average molecular weight of pectin. The degradation kinetics of pectin at different pH values under ultrasound well fitted to a second-order reaction kinetics model. Acoustic cavitation, ß-elimination, and demethylation led to the breakage of glycosidic linkages of side chains and methoxyl groups of pectin, but did not have noticeable influences on the main chain of pectin. The ultrasonic treatment at a high pH led to an apparent change in the rheological characteristics of pectin. Therefore, ultrasonic treatment at various pH values can be developed as a viable means to prepare desirable MP.

Does thyroid autoimmunity affect the reproductive outcome in women with thyroid autoimmunity undergoing assisted reproductive technology?

PROBLEM: Our study aims to investigate whether the anti-thyroperoxidase antibody (TPO-Ab) and TSH level in euthyroid women have any association with reproductive outcomes after the ART cycle. METHODS OF STUDY: A total of 1107 patients who were enrolled in the study were divided into four groups based on serum TSH level and TPO-Ab status: group A, 0.3 ≤ TSH < 2.5 mIU/L and TPO-Ab- ; group B, 0.3 ≤ TSH < 2.5 mIU/L and TPO-Ab+ ; group C, 2.5 ≤ TSH < 4.2 mIU/L, and TPO-Ab- ; and group D, 2.5 ≤ TSH < 4.2 mIU/L, TPO-Ab+ . The differences in ART cycles and pregnancy outcomes were analyzed between study groups. RESULTS: The fertilization rate in group D (73%) was significantly lower than that in groups A (83% P < .001), B (84% P = .001), and C (82% P = .002). The biochemical pregnancy rates of groups B (7%) and D (12%) were significantly higher than those of group A (2%) (P = .028 and P = .017, respectively). TPO-Ab was related to a higher biochemical pregnancy rate (P = .002, OR = 5.311, 95% CI 1.859-15.169) and TSH over 2.5 mIU/L was related to higher ICSI rate (P = .001, OR = 1.759, 95% CI 1.250-2.476) by logistic regression analysis. The receiver operating characteristic (ROC) also verified the results. CONCLUSION: The impacts of TSH ≥ 2.5 mIU/L on the intracytoplasmic sperm injection (ICSI) rate, TSH ≥ 2.5 mIU/L and TPO-Ab+ on the fertilization rate, and TPO-Ab+ on the biochemical pregnancy rate, rather than the effect on abortion, clinical pregnancy, and live birth, were emphasized.

Changes in Concurrent Opioid and Benzodiazepine Use Following a Low-Touch Prescriber Fax Intervention.

BACKGROUND: Concurrent use of opioids and benzodiazepines (COB) can lead to additive respiratory and central nervous system effects, putting patients at increased risk of fatal overdose. In 2016, the Centers for Disease Control and Prevention released an opioid-prescribing guideline recommending against COB, and the Pharmacy Quality Alliance (PQA) endorsed a COB measure in its core opioid set. From May 1, 2017, to December 4, 2017, a California Medicaid plan launched a COB-focused prescriber outreach intervention for members receiving recent opioid and benzodiazepine claims with the intent of decreasing concurrent use. OBJECTIVE: To assess the effect of a prescriber fax intervention by a Medicaid plan on COB. METHODS: Two retrospective analyses were conducted using administrative pharmacy claims data: a comparison of the PQA COB rate among selected California Medicaid plans for 2016 and 2017 and a cohort utilization analysis of members identified for the fax intervention compared with controls. Intervention and control members were matched based on 12 pre-index utilization characteristics. Outcomes assessed included proportion of members with resolution of COB in the post-index period, change in mean number of COB days before and after the index date, and proportion of members with decreased benzodiazepine daily dose after the index date. Analyses were also performed for the subgroups of members with < 30 days of COB and ≥ 30 days of COB in the pre-index period. RESULTS: All California Medicaid plans in the study saw an improvement in the PQA COB rate between 2016 and 2017. In the utilization analysis, 4,182 intervention members were eligible according to study criteria and matched to similar control members. Many differences in medication use existed between the subgroups with < 30 days and ≥ 30 days of COB in the pre-index period, with the latter group consisting of much more chronic, complex users. The intervention cohort had a statistically significant higher proportion of members with complete resolution of COB compared with the control cohort (43.8% vs. 40.0%; P < 0.01), which was also statistically significant for the 2 subgroups. The intervention cohort had a decrease in the mean number of COB days from pre- to post-index periods, but this was only statistically significant for the subgroup with < 30 COB days (-2.5 vs. -1.5; P = 0.0217). No statistically significant differences were detected between cohorts in proportion of members with decreased benzodiazepine dose. CONCLUSIONS: Our analyses demonstrated that this low-touch prescriber fax intervention produced statistically significant improvements in COB outcomes, despite the overall trend of declining COB among the other California Medicaid plans. Low-touch, targeted prescriber outreach can be an inexpensive yet effective tool to affect prescriber behavior, particularly before COB becomes chronic. DISCLOSURES: No outside funding was used to support this study. The authors do not have any financial relationships or potential conflicts of interest relevant to this article to disclose. At the time of conducting this research, all authors were employees of MedImpact Healthcare Systems. The results of this study were presented at the AMCP Managed Care & Specialty Pharmacy Annual Meeting 2019; March 25-28, 2019; San Diego, CA.

Effect of ultrasonic intensity on the conformational changes in citrus pectin under ultrasonic processing.

In this study, the effects of ultrasonic intensity on conformational changes in aqueous citrus pectin solution under ultrasonic processing and its possible transition mechanism were investigated. The results demonstrated that higher ultrasonic intensity (104.7 W/cm2) caused larger alterations in the molecular and conformational parameters of the semiflexible pectin (Mark-Houwink relation exponent a: 0.820, conformational parameter α: 0.607, structural parameter ρ: 2.22) in aqueous solution. Meanwhile, the semiflexible chain of pectin became more flexible (a: 0.804, α: 0.601, ρ: 1.75) at higher ultrasonic intensity in aqueous solution, as was verified by atomic force microscopy. Moreover, conformational changes in pectin from semiflexible chains to flexible chains or even flexible coils (a: 0.791, α: 0.597, ρ: 1.70) could be attributed to the decreased degree of methoxylation and neutral sugars in side chains and the destruction of inter- and intramolecular hydrogen bonds under ultrasonic processing. Therefore, these results have important implications for understanding the ultrasonic modification of pectin.

Quaternized curdlan/pectin polyelectrolyte complexes as biocompatible nanovehicles for curcumin.

In this study, a positively charged quaternized curdlan (Qcurd) was used to fabricate polyelectrolyte complex nanoparticles (PEC NPs) with a negatively charged pectin via electrostatic complexation. Results showed that the Qcurd/pectin PEC NPs prepared with 0.5 mg/mL Qcurd and pectin solutions, 1:2 pectin/Qcurd mass ratio, and pH 4.0 in the absence of NaCl were characterized by a spherical morphology in nanoscale, an average particle size of 68 nm, and good dispersibility in aqueous solutions. Curcumin was encapsulated in the Qcurd/pectin PEC NPs through hydrogen bonding with an encapsulation efficiency of ∼82%, a loading content of 13%, and a pH-dependent controlled release. Curcumin-loaded PEC NPs exhibited a significantly enhanced water solubility, excellent free radical scavenging ability and antioxidant capacity in vitro as compared with those of free curcumin.

Conformational and rheological properties of a quaternary ammonium salt of curdlan.

This study investigated the chain conformation, microstructure, and rheological properties of a quaternary ammonium salt of curdlan (Qcurd) in aqueous medium. Results showed that Qcurd exhibited typical polyelectrolyte behavior in pure water, and the normal viscosity character was observed in 0.1 M NaCl aqueous solution. The weight-average molecular weight (Mw), z-average radius of gyration (Rg), hydrodynamic radius (Rh), conformational parameter (α), characteristic constant (a) and structure parameter (ρ) for Qcurd were determined as 8.08 × 104 g/mol, 26.7 nm, 15.0 nm, 0.54, 0.62, and 1.78, respectively, indicating that Qcurd existed as a flexible chain conformation in 0.1 M NaCl solution, verified by atomic force microscopy. Qcurd exhibited more pronounced shear-thinning behavior at higher concentrations. The flow behavior of Qcurd at different concentrations and temperature was well evaluated by power-law model and Arrhenius equation. Dynamic tests revealed that Qcurd showed oscillatory behaviors between a dilute solution and a weak elastic gel.

Construction, stability, and enhanced antioxidant activity of pectin-decorated selenium nanoparticles.

Selenium nanoparticles (SeNPs) as a new replacement source of other Se forms applied in nutritional supplements have been associated with health-related issues. Pectin (PEC) as a well-known food-grade polysaccharide has been considered as a potential soft template for the preparation and stabilization of SeNPs in aqueous medium. In this study, therefore, PEC was used as a stabilizer and dispersing agent to form well-dispersed and stable SeNPs under a simple redox system of selenite and ascorbic acid. Se/PEC ratios significantly affected the color of the suspension, particle size, and surface morphology of the as-prepared SeNPs in the presence of PEC. PEC-SeNPs with a Se/PEC ratio of 1:2 appeared amorphous and exhibited a well-dispersed and stable spherical structure with an average size of ∼41 nm, which corresponds to the strong hydrogen bonding between the hydroxyl groups of PEC and SeNPs. The PEC-SeNPs (Se/PEC = 1:2) remained highly stable in different acidic solutions for at least 1 month. Small and highly stable PEC-SeNPs (Se/PEC = 1:2) possessed the strongest DPPH radical scavenging ability and antioxidant capacity among the evaluated PEC-SeNPs. They also possessed a low cytotoxic activity against cancer cells (SPCA-1 and HeLa) and normal cells (RWPE-1) in vitro. These findings suggested that pectin as a surface decorator could be effectively used to improve the stability and antioxidant capacity of SeNPs remarkably.

Ultrasound synergized with three-phase partitioning for extraction and separation of Corbicula fluminea polysaccharides and possible relevant mechanisms.

Ultrasound and three-phase partitioning (USTPP) were combined to extract and separate polysaccharides (PS) efficiently from Corbicula fluminea crude extracts. The following optimum experimental conditions were established: 20% (w/v) ammonium sulfate concentration, 1:1 (v/v) t-butanol to crude extract ratio, 180W ultrasonic power, 40kHz frequency, 100% duty cycle, 10min irradiation time, and 35°C. The maximum extraction yield of PS obtained from USTPP was approximately 11.22%, which was higher than the maximum yields from conventional three-phase partitioning (TPP; 9.32%) and ultrasound extraction (USE; 6.05%). The extraction time for USTPP was significantly reduced to 10min from 30 and 60min for TPP and USE methods, respectively. The primary chemical structures of PS collected through the three extraction protocols were basically conserved. The synergistic effects of ultrasound and TPP on PS extraction were also determined. Results revealed that USTPP is an effective technique to extract and separate PS from C. fluminea potentially applied in food, cosmetics, and medicines.

Formation and characterization of polyelectrolyte complex synthesized by chitosan and carboxylic curdlan for 5-fluorouracil delivery.

In this study, negatively charged carboxylic curdlan (Cc) bearing a ß-1,3-polyglucuronic acid structure was employed to fabricate nanosized polyelectrolyte complexes (PECs) with positively charged chitosan (CS) in aqueous solution as potential carriers for 5-fluorouracil (5Fu) delivery. Nanosized CS/Cc PECs were formed by the addition of 0.5mg/mL solutions of CS and Cc with a mixing ratio of 1:1 (w/w) at pH 3.0. Under optimized conditions, the prepared CS/Cc PECs showed spherical morphology with an average size of about 180nm and a zeta potential of around 41mV. The 5Fu drug was incorporated into the nanosized CS/Cc PECs and showed excellent encapsulation efficiency (86.47%) and loading content (10.81%). The drug release data in vitro indicated that the nanosized CS/Cc PECs are promising carriers for the sustained release of 5Fu with an anomalous transport mechanism following the Ritger-Peppas model. Besides, the CS/Cc PECs exhibited low cytotoxic activity against SPCA-1 and HeLa cell lines in vitro. This finding suggested that the development of the nanosized CS/Cc PECs offered great promise as an antitumor drug platform.

Fabrication and stabilization of biocompatible selenium nanoparticles by carboxylic curdlans with various molecular properties.

In this study, carboxylic curdlans (Cur-4, Cur-8, and Cur-24) with different molecular properties and chain conformations were used as stabilizer and capping agent to fabricate stable and water-dispersible selenium nanoparticles (SeNPs). Results showed that molecular properties and chain conformations of carboxylic curdlans remarkably influenced the size, morphology, structure, and stability of SeNPs and the carboxylic curdlan was ligated to SeNPs via OH⋯Se interaction. The as-prepared SeNPs was amorphous and showed homogeneous and monodisperse spherical structure with size of ∼50-90nm. The Cur-8-decorated SeNPs (SeNPs@Cur-8) exhibited smaller particle size (∼56nm) and greater stability than those of the others. The carboxylic curdlan-stabilized SeNPs exhibited excellent antioxidant capacities compared to the control SeNPs. Specifically, SeNPs@Cur-8 with smaller particle size possessed strong antioxidant efficacy. SeNPs@Cur-8 also exhibited low cytotoxic activity against SPCA-1 and HeLa cell lines in vitro.

pH dependent green synthesis of gold nanoparticles by completely C6-carboxylated curdlan under high temperature and various pH conditions.

A C6-carboxylated curdlan (C6-Cc) obtained from 4-acetamido-TEMPO-mediated oxidation of curdlan was used both as a reducing and stabilizing agent for green synthesis of pH-responsive AuNPs, which was carried out by controlling the pH of the C6-Cc solution at a high temperature (100°C). C6-Cc presented a semi-flexible random coil chain in the aqueous medium at pH 5.5 and became more expanded and rigid in alkaline conditions (pH 7.1-12.0), though the primary chemical structure of C6-Cc was virtually unchanged with the pH variation. The AuNPs prepared with C6-Cc at various pHs were characterized by various instrumental measurements. The shapes and sizes of AuNPs were found to be strongly dependent on the pH of the C6-Cc solution. The C6-Cc-decorated AuNPs exhibited a more well-dispersed spherical morphology with smaller particle sizes under alkaline conditions (pH 7.1-12.0). Through this study, a facile, simple, and green method has been demonstrated for preparation of stimuli-sensitive AuNPs using biocompatible polyanionic polysaccharides.

Construction and characterization of nanosized curdlan sulfate/chitosan polyelectrolyte complex toward drug release of zidovudine.

In this study, as-synthesized curdlan sulfate (CRDS) with a flexible random coil chain was selected as a polyanion to construct nanosized polyelectrolyte complexes (PECs) with a polycation, chitosan (CS), via electrostatic interactions. Nanosized PECs were formed simply by mixing CRDS (0.5mg/mL) and CS (0.5mg/mL) at a mass ratio of 2:1 at pH 3.5. The CRDS/CS PECs exhibited a spherical morphology with a Z-average diameter of around 180nm and a zeta potential value of about -38mV. Subsequently, zidovudine (AZT) as a model antiviral drug was successfully encapsulated into the nanosized PECs with a favorable drug loading efficiency. The AZT-loaded PECs presented a controlled and pH-dependent AZT release profile. These findings suggest that the CRDS/CS PECs constructed in the present study can be developed as potential nanocarriers for drug delivery applications.

Three-phase partitioning for efficient extraction and separation of polysaccharides from Corbicula fluminea.

Three-phase partitioning (TPP), which is a simple, efficient, and green bioseparation technique, was used to extract and separate polysaccharides from Corbicula fluminea (CFPS). The optimal parameters with a high yield of 9.32% were as follows: mass fraction of (NH4)2SO4, 20.0% (w/v); amount of t-butanol, 9.8mL; temperature, 35.3°C; extraction time, 30min; and pH 6.0. The purified CFPS after TPP consisted of d-glucose, d-glucosamine, and d-mannose in a molar ratio of 57.1: 5.6:1.0 with high purity (86.5%) and different molecular weights (1311.1 and 41.5-92.8kDa). Amino acid analysis, UV-vis absorption and Fourier transform-infrared spectra indicated that the purified CFPS was a proteoglycan with O-glycosidic bonds. Moreover, the purified CFPS possessed strong free-radical scavenging abilities and antioxidant activities in vitro. The obtained Trolox equivalent antioxidant capacity and ferric-reducing ability of plasma values were 95.01µmol Trolox/g sample and 38.30µmol Fe2+/g sample, respectively.

Biocompatible Polyelectrolyte Complex Nanoparticles from Lactoferrin and Pectin as Potential Vehicles for Antioxidative Curcumin.

Polyelectrolyte complex nanoparticles (PEC NPs) were fabricated via electrostatic interactions between positively charged heat-denatured lactoferrin (LF) particles and negatively charged pectin. The obtained PEC NPs were then utilized as curcumin carriers. PEC NPs were prepared by mixing 1.0 mg/mL solutions of heat-denatured LF and pectin at a mass ratio of 1:1 (w/w) in the absence of NaCl at pH 4.50. PEC NPs that were prepared under optimized conditions were spherical in shape with a particle size of ∼208 nm and zeta potential of ∼-32 mV. Hydrophobic curcumin was successfully encapsulated into LF/pectin PEC NPs with high encapsulation efficiency (∼85.3%) and loading content (∼13.4%). The in vitro controlled release and prominent antioxidant activities of curcumin from LF/pectin PEC NPs were observed. The present work provides a facile and fast method to synthesize nanoscale food-grade delivery systems for the improved water solubility, controlled release, and antioxidant activity of hydrophobic curcumin.

Purification, structural characterization and bioactivity evaluation of a novel proteoglycan produced by Corbicula fluminea.

A novel proteoglycan, named CFPS-11, was isolated from Corbicula fluminea, which is a food source of freshwater bivalve mollusk. CFPS-11 had an average molecular weight of 807.7kDa and consisted of d-glucose and d-glucosamine in a molar ratio of 12.2:1.0. The protein moiety (∼5%) of CFPS-11 was covalently bonded to the polysaccharide chain in O-linkage type through both serine and thereonine residues. The polysaccharide chain of CFPS-11 was composed of (1→4)-α-d-glucopyranosyl and (1→3,6)-α-d-glucopyranosyl residues, which branched at O-6. The branch chain consisted of (1→)-α-d-glucopyranosyl and (1→)-α-d-N-acetylglucosamine residues. CFPS-11 exhibited significant antioxidant activity in a dose-dependent manner and remarkable inhibition activities against α-amylase and α-glucosidase by in vitro assays. These findings indicated that the CFPS-11 from C. fluminea has the potential for development as a health food ingredient.

Is There a Relationship Between Part D Medication Adherence and Part C Intermediate Outcomes Star Ratings Measures?

BACKGROUND: Improvements in the Centers for Medicare & Medicaid Services (CMS) star ratings Part D medication adherence measures may affect performance in Part C intermediate outcome measures for which the Part D targeted medication classes are prescribed. OBJECTIVE: To determine if Part D medication adherence measures are associated with corresponding Part C intermediate outcome measures. METHODS: This was a cross-sectional analysis using the CMS 2015 star ratings report (based on 2013 benefit year plan data) for Medicare contracts. The measures of interest included the Part D adherence measures for diabetes medications, antihypertensive agents, and statins and the Part C intermediate outcome measures for controlled blood sugar, blood pressure, and cholesterol. All Medicare Advantage Prescription Drug (MAPD) contracts with complete data for all Part C and D measures of interest were included. Contracts with ≥ 25% of total enrollment with MA-only benefit were excluded. Linear and logistic regression models were used to assess the association between 2015 Part D adherence measures and Part C intermediate outcome measures (n = 366). The regression models were adjusted for low-income subsidy (LIS) beneficiary enrollment and log-transformed (natural logarithm) total contract enrollment. RESULTS: Bivariate linear regression models demonstrated moderate positive associations between each of the 2015 Part D adherence scores and related 2015 Part C measures that explained 27%-29% (R(2)) of variance. Including LIS and total contract enrollment in the regression models increased the R2 to 30%-36%. The multivariate logistic regression models showed that each percentage point of improvement in the 2015 Part D adherence measures was associated with a 4.13 to 4.69 greater odds of performing in the top quartile in corresponding 2015 Part C measures. CONCLUSIONS: Moderate positive associations were observed between the Part D and Part C scores in the same benefit year. MAPD plans may observe improved Part C intermediate outcome measures with strategies that improve Part D medication adherence measures. DISCLOSURES: This study was conducted by MedImpact Healthcare Systems, San Diego, California, without external funding. All authors are employees of MedImpact Healthcare Systems. Erickson reports advisory board fees from Sanofi and AstraZeneca. Ta, Erickson, and Patel were responsible for study concept and design and data interpretation, with assistance from Qiu. Qiu and Ta took the lead in data collection, assisted by Erickson. Ta wrote the manuscript, which was revised by Erickson and Patel.