Oral formulations for highly lipophilic drugs: Impact of surface decoration on the efficacy of self-emulsifying drug delivery systems.

AIM: To evaluate the impact of the surface decoration of cannabidiol (CBD) loaded self-emulsifying drug delivery systems (SEDDS) on the efficacy of the formulations to cross the various barriers faced by orally administered drugs. METHODS: Polyethylene glycol (PEG)-free polyglycerol (PG)-based SEDDS, mixed zwitterionic phosphatidyl choline (PC)/PEG-containing SEDDS and PEG-based SEDDS were compared regarding stability against lipid degrading enzymes, surface properties, permeation across porcine mucus, cellular uptake and cytocompatibility. RESULTS: SEDDS with a size of about 200 nm with narrow size distributions were developed and loaded with 20-21 % of CBD. For PG containing PEG-free SEDDS increased degradation by lipid degrading enzymes was observed compared to PEG-containing formulations. The surface hydrophobicity of placebo SEDDS increased in the order of PG-based to mixed PC/PEG-based to PEG-based SEDDS. The influence of this surface hydrophobicity was also observed on the ability of the SEDDS to cross the mucus gel layer where highest mucus permeation was achieved for most hydrophobic PEG-based SEDDS. Highest cellular internalization was observed for PEG-based Lumogen Yellow (LY) loaded SEDDS with 92 % in Caco-2 cells compared to only 30 % for mixed PC/PEG-based SEDDS and 1 % for PG-based SEDDS, leading to a 100-fold improvement in cellular uptake for SEDDS having highest surface hydrophobicity. For cytocompatibility all developed placebo SEDDS showed similar results with a cell survival of above 75 % for concentrations below 0.05 % on Caco-2 cells. CONCLUSION: Higher surface hydrophobicity of SEDDS to orally deliver lipophilic drugs as CBD seems to be a promising approach to increase the intracellular drug concentration by an enhanced permeation through the mucus layer and cellular internalization.

Immobilization of Lipase from Thermomyces Lanuginosus and Its Glycerolysis Ability in Diacylglycerol Preparation.

In the glycerolysis process for diacylglycerol (DAG) preparation, free lipases suffer from poor stability and the inability to be reused. To address this, a cost-effective immobilized lipase preparation was developed by cross-linking macroporous resin with poly (ethylene glycol) diglycidyl ether (PEGDGE) followed by lipase adsorption. The selected immobilization conditions were identified as pH 7.0, 35 °C, cross-linking agent concentration 2.0%, cross-linking time 4 h, lipase amount 5 mg/g of support, and adsorption time 4 h. Enzymatic properties of the immobilized lipase were analyzed, revealing enhanced pH stability, thermal stability, storage stability, and operational stability post-immobilization. The conditions for immobilized enzyme-catalyzed glycerolysis to produce DAG were selected, demonstrating the broad applicability of the immobilized lipase. The immobilized lipase catalyzed glycerolysis reactions using various oils as substrates, with DAG content in the products ranging between 35 and 45%, demonstrating broad applicability. Additionally, the changes during the repeated use of the immobilized lipase were characterized, showing that mechanical damage, lipase leakage, and alterations in the secondary structure of the lipase protein contributed to the decline in catalytic activity over time. These findings provide valuable insights for the industrial application of lipase.

Cocos nucifera and glycerine afterwork moisturizers for secondary prevention of hand dermatitis among fabric worker: a randomized, double-blind, cross over trial.

The use of skin barrier-enhancing topical medication is a favorable approach for the treatment of occupational hand dermatitis (OHD). Cocos nucifera or coconut oil is one of the best sources of lipid enriched with laurate acid, and glycerin is a well-known humectant that improves skin hydration. This study is aimed is to evaluate the effectiveness of C. nucifera and glycerin for secondary prevention of OHD among batik (Indonesian traditional fabric) workers. In a randomized, double-blind, crossover trial, the effect of glycerine-C. nucifera cream versus glycerin-only was considered with multiple afterwork applications of moisturizer over a 2-week period on batik workers with OHD. Assessment of trans-epidermal water loss (TEWL), skin capacitance, and a clinical assessment using the Hand Eczema Severity Index (HECSI) were carried out at day 0 and 14. The results show thirty-two batik dyeing and/or rinsing workers were enrolled in the study with mild to moderate OHD. Clinical improvement was demonstrated by 20% decrease in HECSI and TEWL, and 20% increase in skin capacitance. Both moisturizers were equally effective for the secondary prevention of OHD. As a conclusion, glycerine-C. nucifera and glycerin-only cream are equally effective for secondary prevention for OHD among batik worker to reduce the prevalence of hand dermatitis.

Study on encapsulation of Lactobacillus plantarum 23-1 in W/O/W emulsion stabilized by pectin and zein particle complex.

This study was conducted to develop a W/O/W emulsion encapsulated Lactobacillus plantarum 23-1 (LP23-1) to significantly enhance the survival rate of LP23-1 under simulated digestion and storage conditions. The zein particles and pectin formed a complex through electrostatic interaction and hydrogen bonding. When the proportion of zein particles to pectin was 1:1, the emulsifying stability index (ESI) was 304.17 %. Additionally, when the proportion of the internal aqueous phase to the oil phase was 1:9, the polyglycerol polyricinoleate (PGPR) concentration was 5 %, the proportion of primary emulsion to the external aqueous phase was 5:5, the zein particles concentration was 4 %, and the proportion of zein particles to pectin was 1:1, the encapsulation rate was the highest at 96.27 %. Cryo-scanning electron microscopy and fluorescence microscopy confirmed the morphology of W/O/W emulsion and successful encapsulation of LP23-1. Furthermore, compared with free LP23-1, the W/O/W emulsion encapsulation significantly improved the survival rate of LP23-1 to 73.36 % after simulated gastrointestinal digestion and maintained a high survival rate of 78.42 % during the 35-day storage. The W/O/W emulsion was found to effectively improve the survival rate of LP23-1 during simulated digestion and storage, which has implications for the development of probiotic functional foods with elevated survival rates.

Engineering Triphasic Nanocomposite Coatings on Pretreated Mg Substrates for Biomedical Applications.

Biodegradable polymer-based nanocomposite coatings provide multiple advantages to modulate the corrosion resistance and cytocompatibility of magnesium (Mg) alloys for biomedical applications. Biodegradable poly(glycerol sebacate) (PGS) is a promising candidate used for medical implant applications. In this study, we synthesized a new PGS nanocomposite system consisting of hydroxyapatite (HA) and magnesium oxide (MgO) nanoparticles and developed a spray coating process to produce the PGS nanocomposite layer on pretreated Mg substrates, which improved the coating adhesion at the interface and their cytocompatibility with bone marrow derived mesenchymal stem cells (BMSCs). Prior to the spray coating process of polymer-based nanocomposites, the Mg substrates were pretreated in alkaline solutions to enhance the interfacial adhesion strength of the polymer-based nanocomposite coatings. The addition of HA and MgO nanoparticles (nHA and nMgO) to the PGS matrix, as well as the alkaline pretreatment of the Mg substrates, significantly enhanced the interfacial adhesion strength when compared with the PGS coating on the nontreated Mg control. The average BMSC adhesion densities were higher on the PGS/nHA/nMgO coated Mg than the noncoated Mg controls under direct contact conditions. Moreover, the addition of nHA and nMgO to the PGS matrix and coating the nanocomposite onto Mg substrates increased the average BMSC adhesion density when compared with the PGS/nHA/nMgO coated titanium (Ti) and PGS coated Mg controls under direct contact. Therefore, the spray coating process of PGS/nHA/nMgO nanocomposites on Mg substrates or other biodegradable metal substrates could provide a promising surface treatment strategy for biodegradable implant applications.

Ionic co-aggregates based intravenous drug delivery: Evaluation on kinetics and distribution of the drug payloads and nanocarriers.

Surfactants are crucial in formulating poorly soluble drugs but lead to serious side effects due to PEG chains. Novel supra-amphiphiles consisting of fatty acids and choline are developed, which spontaneously form ionic co-aggregates (ICAs) in water and exhibit strong solubilizing capacity. Paclitaxel (PTX) is adopted as a model drug here to evaluate the feasibility of choline oleate-based ICAs in the intravenous delivery of poorly soluble drugs by comparing the kinetics and distribution of payloads and nanocarriers. Choline oleate presents a maximum 10-fold enhancement in solubilizing capacity to PTX than Cremophor EL (CreEL), enabling a one-tenth use level in the formulation. Aggregation-caused quenching probes are utilized to evaluate the kinetics and biodistribution of ICAs or CreEL-based micelles (MCs). A huge gap is found between the pharmacokinetic and particokinetic curves of either nanocarrier, indicating fast leakage. ICAs lead to faster PTX leakage in blood circulation but higher PTX distribution to organs than MCs. MCs present a longer circulation in blood but a slower distribution to organs than ICAs. ICAs do not arise adverse reactions in rats following repeated injections, while MCs cause pathological changes in varying degrees. In conclusion, choline oleate-based ICAs provide an alternative to surfactants in formulating poorly soluble drugs.

Improving an Alternative Glycerol Catabolism Pathway in Yarrowia lipolytica to Enhance Erythritol Production.

Engineering the glycerol-3-phosphate pathway could enhance erythritol production by accelerating glycerol uptake. However, little work has been conducted on the alternative dihydroxyacetone (DHA) pathway in Yarrowia lipolytica. Herein, this route was identified and characterized in Y. lipolytica by metabolomic and transcriptomic analysis. Moreover, the reaction catalyzed by dihydroxyacetone kinase encoded by dak2 was identified as the rate-limiting step. By combining NHEJ-mediated insertion mutagenesis with a push-and-pull strategy, Y. lipolytica strains with high-yield erythritol synthesis from glycerol were obtained. Screening of a library of insertion mutants allows the identification of a mutant with fourfold increased erythritol production. Overexpression of DAK2 and glycerol dehydrogenase GCY3 together with gene encoding transketolase and transaldolase from the nonoxidative part of the pentose phosphate pathway led to a strain with further increased productivity with a titer of 53.1 g/L and a yield 0.56 g/g glycerol, which were 8.1- and 4.2-fold of starting strain.

Automatic characterization of capillary flow profile of liquid samples on µTADs based on capacitance measurement.

Capillary flow profile of liquid samples in porous media is closely related to the important properties of liquid samples, including the viscosity and the surface energy. Therefore, capillary flow profile can be used as an index to differentiate liquid samples with different properties. Fast and automatic characterization of capillary flow profile of liquid samples is necessary. In this work, we develop a portable and economical capacitance acquisition system (CASY) to easily obtain the capillary flow profile of liquid samples on microfluidic thread-based analytical devices (µTADs) by measuring the capacitance during the capillary flow. At first, we validate the accuracy of this method by comparing with the traditional method by video analysis in obtaining the capillary flow profiles in µTADs of cotton threads or glass fiber threads. Then we use it to differentiate liquid samples with different viscosity (mixture of water and glycerol). In addition, capillary flow profile on µTADs with chemical valves (chitosan or sucrose) can also be obtained on this device. Lastly, we show the potential of this device in measurement of hematocrit (HCT) of whole blood samples. This device can be used to catalog liquid biological samples with different properties in point-of-care diagnostics in the near future.

How carbon sources drive cellulose synthesis in two Komagataeibacter xylinus strains.

Bacterial cellulose synthesis from defined media and waste products has attracted increasing interest in the circular economy context for sustainable productions. In this study, a glucose dehydrogenase-deficient Δgdh K2G30 strain of Komagataeibacter xylinus was obtained from the parental wild type through homologous recombination. Both strains were grown in defined substrates and cheese whey as an agri-food waste to assess the effect of gene silencing on bacterial cellulose synthesis and carbon source metabolism. Wild type K2G30 boasted higher bacterial cellulose yields when grown in ethanol-based medium and cheese whey, although showing an overall higher D-gluconic acid synthesis. Conversely, the mutant Δgdh strain preferred D-fructose, D-mannitol, and glycerol to boost bacterial cellulose production, while displaying higher substrate consumption rates and a lower D-gluconic acid synthesis. This study provides an in-depth investigation of two K. xylinus strains, unravelling their suitability for scale-up BC production.

An efficient approach for overproduction of DNA polymerase from Pyrococcus furiosus using an optimized autoinduction system in Escherichia coli.

High fidelity DNA polymerase from Pyrococcus furiosus (Pfupol) is an attractive alternative to the highly popular DNA polymerase from Thermus aquaticus. Because this enzyme is in great demand for biotechnological applications, optimizing Pfupol production is essential to supplying the industry's expanding demand. T7-induced promoter expression in Escherichia coli expression systems is used to express recombinant Pfupol; however, this method is not cost-effective. Here, we have effectively developed an optimized process for the autoinduction approach of Pfupol expression in a defined medium. To better examine Pfupol's activities, its purified fraction was used. A 71 mg/L of pure Pfupol was effectively produced, resulting in a 2.6-fold increase in protein yield when glucose, glycerol, and lactose were added in a defined medium at concentrations of 0.05%, 1%, and 0.6%, respectively, and the condition for production in a 5 L bioreactor was as follow: 200 rpm, 3 vvm, and 10% inoculant. Furthermore, the protein exhibited 1445 U/mg of specific activity when synthesized in its active state. This work presents a high level of Pfupol production, which makes it an economically viable and practically useful approach.

Trait-based study predicts glycerol/diol dehydratases as a key function of the gut microbiota of hindgut-fermenting carnivores.

BACKGROUND: Microbial pdu and cob-cbi-hem gene clusters encode the key enzyme glycerol/diol dehydratase (PduCDE), which mediates the transformation of dietary nutrients glycerol and 1,2-propanediol (1,2-PD) to a variety of metabolites, and enzymes for cobalamin synthesis, a co-factor and shared good of microbial communities. It was the aim of this study to relate pdu as a multipurpose functional trait to environmental conditions and microbial community composition. We collected fecal samples from wild animal species living in captivity with different gut physiology and diet (n = 55, in total 104 samples), determined occurrence and diversity of pdu and cob-cbi-hem using a novel approach combining metagenomics with quantification of metabolic and genetic biomarkers, and conducted in vitro fermentations to test for trait-based activity. RESULTS: Fecal levels of the glycerol transformation product 1,3-propanediol (1,3-PD) were higher in hindgut than foregut fermenters. Gene-based analyses indicated that pduC harboring taxa are common feature of captive wild animal fecal microbiota that occur more frequently and at higher abundance in hindgut fermenters. Phylogenetic analysis of genomes reconstructed from metagenomic sequences identified captive wild animal fecal microbiota as taxonomically rich with a total of 4150 species and > 1800 novel species but pointed at only 56 species that at least partially harbored pdu and cbi-cob-hem. While taxonomic diversity was highest in fecal samples of foregut-fermenting herbivores, higher pduC abundance and higher diversity of pdu/cbi-cob-hem related to higher potential for glycerol and 1,2-PD utilization of the less diverse microbiota of hindgut-fermenting carnivores in vitro. CONCLUSION: Our approach combining metabolite and gene biomarker analysis with metagenomics and phenotypic characterization identified Pdu as a common function of fecal microbiota of captive wild animals shared by few taxa and stratified the potential of fecal microbiota for glycerol/1,2-PD utilization and cobalamin synthesis depending on diet and physiology of the host. This trait-based study suggests that the ability to utilize glycerol/1,2-PD is a key function of hindgut-fermenting carnivores, which does not relate to overall community diversity but links to the potential for cobalamin formation. Video Abstract.

Production of flavor-active compounds and physiological impacts in immobilized Saccharomyces spp. cells during beer fermentation.

Yeast immobilization in beer fermentation has recently regained attention, due to the expansion of the craft beer market and the diversification of styles and flavors. The aim of this study was to evaluate the physiological differences between immobilized and free yeast cells with a focus on flavor-active compounds formation. Three strains of Saccharomyces spp. (SY025, SY067, SY001) were evaluated in both free and immobilized (using a cellulose-based support, referred as ImoYeast) forms during static batch fermentations of 12 °P malt extract. Immobilized cells showed higher glycerol (SY025, 40%; SY067, 53%; SY001, 19%) and biomass (SY025, 67%; SY067, 78%; SY001, 56%) yields than free cells. Conversely, free cells presented higher ethanol yield (SY025, 9%; SY067, 9%; SY001, 13%). Flavor-active compounds production exhibited significant alterations between immobilized and free cells systems, for all strains tested. Finally, a central composite design with varying initial biomass (X0) and substrate (S0) concentrations was conducted using strain SY025, which can be helpful to modulate the formation of one or more flavor-active compounds. In conclusion, yeast immobilization in the evaluated support resulted in flavor alterations that can be exploited to produce different beer styles.

Efficacy assessment of different cryoprotectants for preserving the viability of Enterobacterales strains at - 20 °C.

The preservation of microorganisms is pivotal in microbiological practice. Currently, cryopreservation is assumed to be an effective and inexpensive approach for the storage of microorganisms, including bacteria. The key point of cryopreservation is optimal cryoprotectant selection. In the present study, different cryoprotectant compositions were tested for long-term storage of 15 Enterobacterales bacterial strains at - 20 °C. The survival rates of the bacterial strains were evaluated in four different cryoprotectant solutions containing 70% glycerin only (cryoprotectants 1 and 4), 10% dimethyl sulfoxide (DMSO) with 70% glycerin (cryoprotectant 2), and 10% DMSO (cryoprotectant 3). In addition, cryoprotectants 1 and 2 contained peptone and yeast extract as nutritional supplements. The general survival rates of the bacterial strains were evaluated after 12 months of storage. After 12 months, the survival rates of the different cryoprotectants were as follows: cryoprotectant 1-88.87%; cryoprotectant 2-84.85%; cryoprotectant 3-83.50%; and cryoprotectant 4-44.81%. Thus, the composition of cryoprotectant 1 (70% glycerin with nutrient supplements) was optimal for preserving 15 tested strains of the order Enterobacterales. Despite these findings, the biochemical properties of the tested strains changed after cryopreservation for 12 months in the presence of 1 or 3 cryoprotectants. Alterations in the biochemical profile could be related to changes in environmental conditions and cold adaptation. We assume that the composition of cryoprotectant 1 can be optimal for storing the order Enterobacterales at - 20 °C. However, further investigations are needed to elucidate the problem of cryopreservation and to support our assumption.

Engineering Escherichia coli for D-allulose biosynthesis from glycerol.

D-allulose, a naturally occurring monosaccharide, is present in small quantities in nature. It is considered a valuable low-calorie sweetener due to its low absorption in the digestive tract and zero energy for growth. Most of the recent efforts to produce D-allulose have focused on in vitro enzyme catalysis. However, microbial fermentation is emerging as a promising alternative that offers the advantage of combining enzyme manufacturing and product synthesis within a single bioreactor. Here, a novel approach was proposed for the efficient biosynthesis of D-allulose from glycerol using metabolically engineered Escherichia coli. FbaA, Fbp, AlsE, and A6PP were used to construct the D-allulose synthesis pathway. Subsequently, PfkA, PfkB, and Pgi were disrupted to block the entry of the intermediate fructose-6-phosphate (F6P) into the Embden-Meyerhof-Parnas (EMP) and pentose phosphate (PP) pathways. Additionally, GalE and FryA were inactivated to reduce D-allulose consumption by the cells. Finally, a fed-batch fermentation process was implemented to optimize the performance of the cell factory. As a result, the titer of D-allulose reached 7.02 g/L with a maximum yield of 0.287 g/g.

Viscoelastic and Birefringence Relaxation of Individualized Cellulose Nanofibers in the Dilute and Semidilute Regions.

Viscoelastic relaxation mechanisms of individualized cellulose nanofibers (iCNFs) dispersed in glycerol in the dilute and semidilute regions were investigated by linear viscoelastic and dynamic birefringence measurements. The birefringence relaxation of the iCNFs was described by the orientational and curvature modes of an existing viscoelastic theory for ideal semiflexible polymers (Shankar-Pasquali-Morse theory). However, the Shankar-Pasquali-Morse theory could not fully describe the iCNF viscoelastic relaxation at high frequencies. Considering the results for birefringence relaxation, the experimental tension mode of the iCNFs was evaluated to be higher than the theoretical value. These results show that the viscoelastic relaxations of the iCNFs are different from those of ideal semiflexible polymers, in contrast to cellulose nanocrystals (CNCs). As the iCNF concentration increased, the orientational mode dramatically slowed, which was more drastic than other semiflexible polymers, including CNCs. This anomalous behavior is likely due to the nonideal nature of iCNFs.

Carbon Quantum Dot-Functionalized Dermis-Derived Transparent Electronic Skin for Multimodal Human Motion Signal Monitoring and Construction of Self-Powered Triboelectric Nanogenerator.

Electronic skin (e-skin) is considered as a highly promising interface for human-computer interaction systems and wearable electronic devices. Through elaborate design and assembly of various materials, it possesses multiple characteristics similar to human skin, including remarkable flexibility, stretchability, sensitivity to temperature and humidity, biocompatibility, and efficient interfacial ion/electron transport capabilities. Here, we innovatively integrate multifunctional carbon quantum dots (CQDs), which exhibit conductivity, antibacterial properties, ultraviolet absorption, and fluorescence emission, with poly(acrylic acid) and glycerin (Gly) into a three-dimensional network structure of natural goatskin collagen fibers. Through a top-down design strategy enhanced by hydrogen bond reconstruction, we successfully fabricated a novel transparent e-skin (PAC-eSkin). This e-skin exhibited significant tensile properties (4.94 MPa of tensile strength and 263.42% of a maximum breaking elongation), while also possessing Young's modulus similar to human skin (2.32 MPa). It is noteworthy that the functionalized CQDs used was derived from discarded goat hair, and the addition of Gly gave PAC-eSkin excellent antifreezing and moisturizing properties. Due to the presence of ultrasmall CQDs, which creates efficient ion/electron transport channels within PAC-eSkin, it could rapidly sense human motion and physiological signals (with a gauge factor (GF) of 1.88). Furthermore, PAC-eSkin had the potential to replace traditional electrode patches for real-time monitoring of electrocardiogram, electromyogram, and electrooculogram signals, with a higher SNR (signal-to-noise ratio) of 25.1 dB. Additionally, the customizable size and shape of PAC-eSkin offer vast possibilities for the construction of single-electrode triboelectric nanogenerator systems. We have reason to believe that the design and development of this transparent e-skin based on CQDs-functionalized dermal collagen matrices can pave a new way for innovations in human-computer interaction interfaces and their sensing application in diverse scenarios.

Glycerol metabolism contributes to competition by oral streptococci through production of hydrogen peroxide.

As a biological byproduct from both humans and microbes, glycerol's contribution to microbial homeostasis in the oral cavity remains understudied. In this study, we examined glycerol metabolism by Streptococcus sanguinis, a commensal associated with oral health. Genetic mutants of glucose-PTS enzyme II (manL), glycerol metabolism (glp and dha pathways), and transcriptional regulators were characterized with regard to glycerol catabolism, growth, production of hydrogen peroxide (H2O2), transcription, and competition with Streptococcus mutans. Biochemical assays identified the glp pathway as a novel source for H2O2 production by S. sanguinis that is independent of pyruvate oxidase (SpxB). Genetic analysis indicated that the glp pathway requires glycerol and a transcriptional regulator, GlpR, for expression and is negatively regulated by PTS, but not the catabolite control protein, CcpA. Conversely, deletion of either manL or ccpA increased the expression of spxB and a second, H2O2-non-producing glycerol metabolic pathway (dha), indicative of a mode of regulation consistent with conventional carbon catabolite repression (CCR). In a plate-based antagonism assay and competition assays performed with planktonic and biofilm-grown cells, glycerol greatly benefited the competitive fitness of S. sanguinis against S. mutans. The glp pathway appears to be conserved in several commensal streptococci and actively expressed in caries-free plaque samples. Our study suggests that glycerol metabolism plays a more significant role in the ecology of the oral cavity than previously understood. Commensal streptococci, though not able to use glycerol as a sole carbohydrate source for growth, benefit from the catabolism of glycerol through production of both ATP and H2O2. IMPORTANCE: Glycerol is an abundant carbohydrate in the oral cavity. However, little is understood regarding the metabolism of glycerol by commensal streptococci, some of the most abundant oral bacteria. This was in part because most streptococci cannot grow on glycerol as the sole carbon source. In this study, we show that Streptococcus sanguinis, a commensal associated with dental health, can degrade glycerol for persistence and competition through two pathways, one of which generates hydrogen peroxide at levels capable of inhibiting Streptococcus mutans. Preliminary studies suggest that several additional commensal streptococci are also able to catabolize glycerol, and glycerol-related genes are actively expressed in human dental plaque samples. Our findings reveal the potential of glycerol to significantly impact microbial homeostasis, which warrants further exploration.

Tailoring solid-state DNP methods to the study of α-synuclein LLPS.

Dynamic Nuclear Polarization (DNP) is a technique that leverages the quantum sensing capability of electron spins to enhance the sensitivity of nuclear magnetic resonance (NMR) signals, especially for insensitive samples. Glassing agents play a crucial role in the DNP process by facilitating the transfer of polarization from the unpaired electron spins to the nuclear spins along with cryoprotection of biomolecules. DNPjuice comprising of glycerol-d8/D2O/H2O has been extensively used for this purpose over the past two decades. Polyethylene glycol (PEG), also used as a cryoprotectant, is often used as a crowding agent in experimental setups to mimic cellular conditions, particularly the invitro preparation of liquid-liquid phase separated (LLPS) condensates. In this study, we investigate the efficacy of PEG as an alternative to glycerol in the DNP juice, critical for signal enhancement. The modified DNP matrix leads to high DNP enhancement which enables direct study of LLPS condensates by solid-state DNP methods without adding any external constituents. An indirect advantage of employing PEG is that the PEG signals appear at ∼72.5 ppm and are relatively well-separated from the aliphatic region of the protein spectra. Large cross-effect DNP enhancement is attained for 13C-glycine by employing the PEG-water mixture as a glassing agent and ASYMPOL-POK as the state-of-art polarizing agent, without any deuteration. The DNP enhancement and the buildup rates are similar to results obtained with DNP juice, conforming to that PEG serves as a good candidate for both inducing crowding and glassing agent in the study of LLPS.