Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Lentils ( Lens culinaris ; family: Fabaceae) are increasingly recognized as functional legumes with potential benefits for gut health because they provide bioactive peptides, resistant starch, and polyphenol-rich fractions within a shared food matrix. However, most existing studies have focused on individual lentil-derived compounds, and their matrix-dependent complementary interactions during digestion and fermentation remain insufficiently resolved. This review synthesizes current evidence on lentil-derived peptides, resistant starch, and polyphenols, with particular emphasis on their matrix-dependent complementary relationships, digestion-dependent transformation, microbial co-metabolism, and implications for intestinal barrier function. During gastrointestinal digestion and colonic fermentation, lentil proteins, resistant starch, and phenolic compounds undergo sequential transformation, yielding bioactive peptides, fermentable substrates, short-chain fatty acids (SCFAs), and phenolic metabolites that may collectively influence microbial composition and metabolic activity. Emerging evidence suggests that these interconnected processes may support gut health through microbiota–host crosstalk by modulating tight junction-related markers, reducing intestinal permeability, and maintaining epithelial homeostasis. Mechanistically, these effects have been associated with SCFA-mediated G protein-coupled receptor (GPCR) signaling, suppression of TLR4–NF-κB/MAPK inflammatory cascades, and activation of Keap1–Nrf2 antioxidant defenses, thereby attenuating oxidative stress and pro-inflammatory responses. Current evidence is more consistent with matrix-dependent complementary or convergent actions than with demonstrated synergy. At present, phenolic-rich fractions provide clear pathway-level evidence, whereas fermentation-linked carbohydrate effects are more strongly supported by microbiota- and in vivo-associated outcomes, and protein- or peptide-related mechanisms remain comparatively underdefined. Nevertheless, the evidence base remains limited by the scarcity of integrated studies, well-controlled human intervention trials, and factorial experimental designs capable of distinguishing complementary, additive, and truly synergistic effects among lentil bioactives. This review therefore highlights the need to move from describing coexisting beneficial effects toward formally testing interaction effects within physiologically relevant lentil matrices.

Candidates:
A. Lentils ( Lens culinaris ; family: Fabaceae) are increasingly recognized as functional legumes with potential benefits for gut health because they provide bioactive peptides, resistant starch, and polyphenol-rich fractions within a shared food matrix.
B. During gastrointestinal digestion and colonic fermentation, lentil proteins, resistant starch, and phenolic compounds undergo sequential transformation, yielding bioactive peptides, fermentable substrates, short-chain fatty acids (SCFAs), and phenolic metabolites that may collectively influence microbial composition and metabolic activity.
C. The evidence does not state that however, most existing studies have focused on individual lentil-derived compounds, and their matrix-dependent complementary interactions during digestion and fermentation remain insufficiently resolved.
D. This review synthesizes current evidence on lentil-derived peptides, resistant starch, and polyphenols, with particular emphasis on their matrix-dependent complementary relationships, digestion-dependent transformation, microbial co-metabolism, and implications for intestinal barrier function.
E. Lentils ( Lens culinaris ; family: Fabaceae) are not increasingly recognized as functional legumes with potential benefits for gut health because they provide bioactive peptides, resistant starch, and polyphenol-rich fractions within a shared food matrix.
F. However, most existing studies have focused on individual lentil-derived compounds, and their matrix-dependent complementary interactions during digestion and fermentation remain insufficiently resolved.
G. The evidence does not state that during gastrointestinal digestion and colonic fermentation, lentil proteins, resistant starch, and phenolic compounds undergo sequential transformation, yielding bioactive peptides, fermentable substrates, short-chain fatty acids (SCFAs), and phenolic metabolites that may collectively influence microbial composition and metabolic activity.
H. The evidence does not state that this review synthesizes current evidence on lentil-derived peptides, resistant starch, and polyphenols, with particular emphasis on their matrix-dependent complementary relationships, digestion-dependent transformation, microbial co-metabolism, and implications for intestinal barrier function.


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
L-theanine is a characteristic non-proteinogenic amino acid found in tea leaves and has attracted considerable attention because of its diverse physiological activity and broad application prospects. γ-glutamyl transpeptidase (GGT) can catalyze the synthesis of L-theanine from L-glutamine and ethylamine without ATP consumption, highlighting its advantages for enzymatic production. In this study, a complete process was established for L-theanine production. Through screening of single and dual promoters, the optimal expression combination, PyxiE-PspoVG, was identified. Furthermore, by integrating the dal selection marker, an antibiotic-free engineered strain was developed. After flask-level optimization, the GGT activity reached 27.32 U/mL and further increased to 127.37 U/mL in 3 L fed-batch fermentation. Using the fermentation broth as the biocatalyst, fed-batch conversion of 0.6 M L-glutamine and 2 M ethylamine yielded 0.52 M (91.44 g/L) L-theanine within 24 h. Further integration of ceramic membrane filtration, ultrafiltration, nanofiltration, electrodialysis, activated-carbon decolorization and ethanol crystallization afforded a final product purity of 95.6%. This study offers a useful reference for large-scale L-theanine production.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Triple-negative breast cancer (TNBC) remains a significant challenge in oncology, contributing to a significant portion of cancer-related deaths among women. Current therapeutic options, including chemotherapy, surgery, radiation, and hormonal targeting therapies, exhibit limited efficacy, necessitating the exploration of innovative treatment modalities. The emergence of drug resistance and the persistence of cancer stem cells (CSCs) further emphasize the urgent need for novel therapeutic strategies. In this context, natural killer cell-derived extracellular vesicles (NK-EVs) have emerged as a promising cell-free therapeutic approach that exhibits high tumor infiltration and cytotoxicity against cancer cells and CSCs. This study aims to investigate the efficacy of NK-EVs as a therapeutic strategy for TNBC using various clinically relevant models, including patient-derived xenografts. Pathway analysis suggests strong activation of apoptosis via canonical caspase activation, as well as necrosis, thereby confirming the important cytotoxic effect of NK-EVs. Interestingly, NK-EVs were also found to suppress TNBC CSCs by disrupting their functionality and viability, and NK-EV treatment increased the expression of apoptosis markers in both CSCs and non-CSCs. By elucidating the therapeutic efficacy and translational potential of NK-EV-based interventions in TNBC, these findings offer critical insights for the development of future immunotherapeutic strategies against this aggressive subtype of breast cancer.

Candidates:
A. The evidence does not state that in this context, natural killer cell-derived extracellular vesicles (NK-EVs) have emerged as a promising cell-free therapeutic approach that exhibits high tumor infiltration and cytotoxicity against cancer cells and CSCs.
B. Current therapeutic options, including chemotherapy, surgery, radiation, and hormonal targeting therapies, exhibit limited efficacy, necessitating the exploration of innovative treatment modalities.
C. In this context, natural killer cell-derived extracellular vesicles (NK-EVs) have emerged as a promising cell-free therapeutic approach that exhibits high tumor infiltration and cytotoxicity against cancer cells and CSCs.
D. Triple-negative breast cancer (TNBC) remains a significant challenge in oncology, contributing to a significant portion of cancer-related deaths among women.
E. The evidence does not state that triple-negative breast cancer (TNBC) remains a significant challenge in oncology, contributing to a significant portion of cancer-related deaths among women.
F. The emergence of drug resistance and the persistence of cancer stem cells (CSCs) further emphasize the urgent need for novel therapeutic strategies.
G. The evidence does not state that current therapeutic options, including chemotherapy, surgery, radiation, and hormonal targeting therapies, exhibit limited efficacy, necessitating the exploration of innovative treatment modalities.
H. The evidence does not state that the emergence of drug resistance and the persistence of cancer stem cells (CSCs) further emphasize the urgent need for novel therapeutic strategies.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Monoclonal antibody N -glycosylation is a critical quality attribute influencing therapeutic safety and efficacy, and is strongly influenced by bioprocess design. NISTCHO, a publicly available Chinese hamster ovary producer cell line, is increasingly encouraged for use as a reference system. However, the impact of feeding strategies on cellular performance and N -glycosylation has not been assessed. Here, we applied multivariate analysis of compositional N -glycan data to assess how feeding strategies influence N -glycan composition of cNISTmAb. We varied feeding strategies in frequency, glucose supply, and galactose/manganese supplementation. Feeding frequency had minimal impact on quality attributes but strongly affected culture performance, with every-other-day feeding improving titers and cell-specific productivity. High glucose availability supported growth and productivity. Low glucose strategies reduced titers and shifted N -glycosylation towards non-galactosylated and fucosylated species, despite lactate accumulation remaining within favorable ranges. Galactose and manganese consistently increased antibody galactosylation, with galactose additionally serving as an auxiliary carbon source, extending cell viability. Importantly, mAb glycation remained stable across all feeding strategies at harvest. Overall, these results demonstrate that feed composition and timing can be used to tune both cellular performance and mAb glycosylation, establishing NISTCHO as a robust benchmark for standardized process-quality studies. Subject terms: Biochemistry, Biological techniques, Biotechnology

Candidates:
A. The evidence does not state that here, we applied multivariate analysis of compositional N -glycan data to assess how feeding strategies influence N -glycan composition of cNISTmAb.
B. Here, we applied multivariate analysis of compositional N -glycan data to assess how feeding strategies influence N -glycan composition of cNISTmAb.
C. Monoclonal antibody N -glycosylation is a critical quality attribute influencing therapeutic safety and efficacy, and is strongly influenced by bioprocess design.
D. The evidence does not state that however, the impact of feeding strategies on cellular performance and N -glycosylation has not been assessed.
E. NISTCHO, a publicly available Chinese hamster ovary producer cell line, is not increasingly encouraged for use as a reference system.
F. NISTCHO, a publicly available Chinese hamster ovary producer cell line, is increasingly encouraged for use as a reference system.
G. Monoclonal antibody N -glycosylation is not a critical quality attribute influencing therapeutic safety and efficacy, and is strongly influenced by bioprocess design.
H. However, the impact of feeding strategies on cellular performance and N -glycosylation has not been assessed.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["120 g/l", "121 g/l", "20 g/l", "21 g/l"]

Evidence:
The filamentous fungus Thermothelomyces heterothallica C1 has been developed into a highly productive protein production system for heterologous proteins like antibodies and vaccine candidates. While it is capable of secreting over 120 g/l of its native enzymes, monoclonal antibodies (mAbs) have been produced at titers exceeding 20 g/l, and strains engineered to produce human-type N-glycan structures have been developed. However, significant variability in mAb productivity and reduced production levels in glycoengineered strains limit the use of C1 as a widespread production host in the pharmaceutical industry. To address these issues, transcriptome analysis was conducted on strains producing five different mAbs with varying production efficiencies, as well as on mAb-producing strains with native and glycoengineered N-glycans. Genes related to protein folding and secretion, which are regulated in response to mAb production and glycoengineering, were over-expressed in a glycoengineered mAb producing C1 strain. In addition, genes identified based on previously described functions in the secretory pathway, including counterparts of human origin, were included in the study. Transcriptome analysis revealed that the mAb heavy and light chains were among the most abundantly expressed transcripts, indicating that production bottlenecks occur after transcription. Genes associated with protein folding, quality control, glycosylation, and transport within the secretory pathway were upregulated in the mAb-producing strains. This upregulation was more pronounced in strains with low mAb yields and in glycoengineered strains. The over-expression of 10 genes ( bet1 , dnaj -type gene, dpm1 , ero1 , erv46 , human calreticulin, human cypb , human mzb1 , pmr1 , and UDP-galactose transporter), each playing distinct roles in the secretory pathway, enhanced mAb production in glycoengineered C1 strains from 1.5- to 2.5-fold. Through a comprehensive analysis of transcriptome data from C1 strains producing various monoclonal antibodies (mAbs) and an extensive literature search, several factors related to protein folding and secretion were identified as potential targets for enhancing mAb production. The over-expression of some of these genes in glycoengineered C1 strains led to improvement in mAb production, with some genes enhancing mAb yields by 2.5-fold. The online version contains supplementary material available at 10.1186/s12934-026-02989-w.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Thermothelomyces thermophilus is a filamentous fungus isolated from self-heating compost. Unlike most of the fungal kingdom, this species exhibits a growth optimum at 45°C and is intolerant of temperatures below 30°C. To investigate genetic contributors to temperature-dependent fitness in this system, we implemented a large-scale insertional mutagenesis approach. We generated thousands of T. thermophilus mutants and cultured them at temperature extremes in a standard medium. Phenotyping-by-sequencing identified dozens of disrupted loci representing candidate determinants of thermophilic life history, including several annotated in metal transport. We then validated a subset of screen hits with a directed, single-gene knockout paradigm. The results revealed a temperature-dependent regulatory logic for germination, the developmental decision by which a fungal spore initiates growth. Surprisingly, most mutants germinated far better at 50°C than the wild type in a standard medium and showed markedly slower germination at lower temperatures, consistent with altered germination regulation rather than enhanced intrinsic heat tolerance. We hypothesized that T. thermophilus has evolved sophisticated regulatory machinery to block germination at high temperatures unless environmental conditions are favorable. As a proof of concept, we surveyed media conditions and established that elevated zinc dampened germination of wild-type T. thermophilus at 50°C but promoted it at lower temperatures; mutation experiments made clear that such sensitivity was mediated in part by the zinc transporter zip . We interpret these results under a model in which T. thermophilus integrates temperature and nutrient availability to control the transition from spore dormancy to vegetative growth, a developmental decision that shapes fitness outcomes across temperatures. Fungal thermophiles thrive at temperatures that represent the upper limits of eukaryotic life. The regulatory and developmental mechanisms that shape their temperature-dependent fitness remain poorly understood. In this work, we elucidate how Thermothelomyces thermophilus integrates temperature cues with other environmental inputs during germination, a key life-cycle stage for dispersal. Our findings highlight germination regulation as an important contributor to fitness at elevated temperatures in a thermophilic eukaryote. These insights are of basic biological interest and provide a foundation for rational strategies to modulate temperature-dependent performance in industrial strains, with applications for high-temperature bioprocessing.

Candidates:
A. Thermothelomyces thermophilus is a filamentous fungus isolated from self-heating compost.
B. Unlike most of the fungal kingdom, this species exhibits a growth optimum at 45°C and is intolerant of temperatures below 30°C.
C. To investigate genetic contributors to temperature-dependent fitness in this system, we implemented a large-scale insertional mutagenesis approach.
D. The evidence does not state that to investigate genetic contributors to temperature-dependent fitness in this system, we implemented a large-scale insertional mutagenesis approach.
E. thermophilus mutants and cultured them at temperature extremes in a standard medium.
F. The evidence does not state that thermophilus mutants and cultured them at temperature extremes in a standard medium.
G. Thermothelomyces thermophilus is not a filamentous fungus isolated from self-heating compost.
H. Unlike most of the fungal kingdom, this species exhibits a growth optimum at 46°C and is intolerant of temperatures below 30°C.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Transferrin is one of the major soluble serum proteins and is responsible for iron transport. Industrially, it is significant as a component of mammalian cell culture media, where a safe and stable supply is necessary. However, because transferrin is a glycoprotein containing 19 disulfide bonds, it is difficult to produce as a recombinant protein in bacteria, and at present it is mainly sourced from animals. In glycoprotein production, stability of the N -glycan profile is crucial, as glycans play important roles in diverse biological processes and influence the efficacy of glycoproteins. In this study, we aimed to produce recombinant human transferrin (rhTF) with stable N -glycan profiles. We generated transgenic rice calli expressing human TF (hTF) as a secretory glycosylated protein. rhTF was successfully produced as a soluble protein in the liquid culture medium of transgenic rice calli and subsequently purified. We confirmed that rhTF contained two plant-specific N -glycans and that these profiles were consistent across production batches. Purified rhTFs promoted the proliferation of cultured animal cells and human iPS cells, similar to serum-derived transferrin. Our results demonstrate new possibilities for producing recombinant glycoproteins with stable N -glycan profiles using a plant cell culture-based secretory protein expression system. Keywords: N -glycan profile, recombinant protein, rice, transferrin

Candidates:
A. Transferrin is one of the major soluble serum proteins and is responsible for iron transport.
B. Industrially, it is significant as a component of mammalian cell culture media, where a safe and stable supply is necessary.
C. Transferrin is not one of the major soluble serum proteins and is responsible for iron transport.
D. However, because transferrin is a glycoprotein containing 19 disulfide bonds, it is difficult to produce as a recombinant protein in bacteria, and at present it is mainly sourced from animals.
E. In glycoprotein production, stability of the N -glycan profile is crucial, as glycans play important roles in diverse biological processes and influence the efficacy of glycoproteins.
F. However, because transferrin is a glycoprotein containing 20 disulfide bonds, it is difficult to produce as a recombinant protein in bacteria, and at present it is mainly sourced from animals.
G. In glycoprotein production, stability of the N -glycan profile is not crucial, as glycans play important roles in diverse biological processes and influence the efficacy of glycoproteins.
H. Industrially, it is not significant as a component of mammalian cell culture media, where a safe and stable supply is necessary.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Direct manufacturing of alloys through casting is economical, user-friendly and conducive to near-net-shape processing, and therefore of great application interest. However, as-cast alloys are rarely used directly because, without post-cast thermomechanical treatments, the solidification product is susceptible to coarse grains, compositional segregation and particularly unsatisfying precipitates, which tend towards inadequate strength and brittleness. These ubiquitous disadvantages have, for a long time, seriously hindered practical applications. Here, through screening by using thermodynamic calculations and calorimetric monitoring, we successfully landed non-equiatomic NiCoCrAlTaZrB complex-concentrated alloys (CCAs) to realize in situ micro-segregation-induced size-gradient L1 2 nanoprecipitates increasing from dendritic to interdendritic regions. Consequently, without any post-cast treatment, prolific in situ nanoprecipitation strengthening renders ultrahigh as-cast yield strength at the gigapascal (GPa) level, while multiscale chemical/structural segregation heterogeneities reinforced by gradient nanoprecipitates result in progressive and persistent hetero-deformation within dendrites, thereby sustaining a high strain-hardening rate of ∼3 GPa and uniform tensile elongation of up to ∼32%. Such robust as-cast strength−ductility synergy not only exceeds all previous as-cast CCAs, but also outperforms all commercial alloys that have already been optimized via post-cast treatments. This advance through exploiting in situ size-gradient nanoprecipitation accomplishes a goal that is more practical than the common pursuit for property records, which all require downstream (multistep and/or subtractive) processing that increases production time and expenses, and even raises severe environmental concerns.

Candidates:
A. Here, through screening by using thermodynamic calculations and calorimetric monitoring, we successfully landed non-equiatomic NiCoCrAlTaZrB complex-concentrated alloys (CCAs) to realize in situ micro-segregation-induced size-gradient L2 2 nanoprecipitates increasing from dendritic to interdendritic regions.
B. The evidence does not state that these ubiquitous disadvantages have, for a long time, seriously hindered practical applications.
C. Direct manufacturing of alloys through casting is not economical, user-friendly and conducive to near-net-shape processing, and therefore of great application interest.
D. Direct manufacturing of alloys through casting is economical, user-friendly and conducive to near-net-shape processing, and therefore of great application interest.
E. However, as-cast alloys are rarely used directly because, without post-cast thermomechanical treatments, the solidification product is not susceptible to coarse grains, compositional segregation and particularly unsatisfying precipitates, which tend towards inadequate strength and brittleness.
F. These ubiquitous disadvantages have, for a long time, seriously hindered practical applications.
G. However, as-cast alloys are rarely used directly because, without post-cast thermomechanical treatments, the solidification product is susceptible to coarse grains, compositional segregation and particularly unsatisfying precipitates, which tend towards inadequate strength and brittleness.
H. Here, through screening by using thermodynamic calculations and calorimetric monitoring, we successfully landed non-equiatomic NiCoCrAlTaZrB complex-concentrated alloys (CCAs) to realize in situ micro-segregation-induced size-gradient L1 2 nanoprecipitates increasing from dendritic to interdendritic regions.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["501 hours", "500 hours", "3.7 g/L", "2.7 g/L"]

Evidence:
Halogenation enhances the stability and function of pharmaceuticals, biomaterials, and industrial compounds. However, chemical halogenation of molecules and peptides can lack stereoselectivity and require the use of toxic chemicals. Although enzymatic halogenation can improve selectivity and reduce environmental impact, current halogenases are inefficient and insoluble, leading to low yields that limit their applications. Here, we develop RebH Evo4 , a soluble and highly active tryptophan halogenase, containing 12 mutations that confer 37-fold and 44-fold increases in 7-chloro- and 7-bromotryptophan production respectively, in vivo. To create RebH Evo4 , we devise an aminoacyl-tRNA synthetase-based halogenase biosensor and conduct over 500 hours of phage-assisted continuous evolution (PACE). Use of RebH Evo4 in a bioreactor results in the production of 2.7 g/L of halogenated tryptophan. When coupled with a downstream enzyme, RebH Evo4 allows 36-fold increased yields of halogenated tryptamines compared to the wild-type enzyme. Additionally, RebH Evo4 enables efficient production of genetically encoded antimicrobial halogenated peptides. The efficient, site-specific halogenation enabled by our evolved halogenase will accelerate sustainable biomanufacturing of halogenated drugs.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Indigo, a ubiquitous vat dye in denim manufacturing, is characterized by its exceptionally low aqueous solubility, necessitating chemical reduction to its leuco form using hazardous agents such as sodium dithionite. This conventional process yields sulfite, sulfate, and sulfide byproducts, leading to significant environmental and toxicological concerns. To address these limitations, this study synthesized 1:1 molar ratio inclusion complexes of indigo with β-cyclodextrin (β-CD) and hydroxypropyl-β-cyclodextrin (HP-β-CD) to enhance solubility and stability without chemical intervention. The formation of these complexes was rigorously validated through UV–Vis, FT-IR, PXRD, TEM, and 1 H-NMR spectroscopy. Notably, the aqueous solubility of indigo was enhanced 3.58-fold and 5.55-fold for the β-CD and HP-β-CD complexes, respectively. Furthermore, both complexes demonstrated superior thermal and photostability, with HP-β-CD exhibiting the most pronounced effects. This cyclodextrin-assisted solubilization was successfully extended to naturally extracted Jeju Indigo, underscoring its broad applicability. Our findings suggest that cyclodextrin-based encapsulation offers a sustainable and effective alternative to conventional reduction-oxidation dyeing processes. The online version contains supplementary material available at 10.1186/s40643-026-01065-w. Keywords: Indigo, β‑cyclodextrin, Hydroxypropyl‑β‑cyclodextrin, Inclusion complex, Solubility enhancement, Reducing agent-free dyeing

Candidates:
A. The formation of these complexes was rigorously validated through UV–Vis, FT-IR, PXRD, TEM, and 1 H-NMR spectroscopy.
B. To address these limitations, this study synthesized 2:1 molar ratio inclusion complexes of indigo with β-cyclodextrin (β-CD) and hydroxypropyl-β-cyclodextrin (HP-β-CD) to enhance solubility and stability without chemical intervention.
C. Indigo, a ubiquitous vat dye in denim manufacturing, is not characterized by its exceptionally low aqueous solubility, necessitating chemical reduction to its leuco form using hazardous agents such as sodium dithionite.
D. Indigo, a ubiquitous vat dye in denim manufacturing, is characterized by its exceptionally low aqueous solubility, necessitating chemical reduction to its leuco form using hazardous agents such as sodium dithionite.
E. To address these limitations, this study synthesized 1:1 molar ratio inclusion complexes of indigo with β-cyclodextrin (β-CD) and hydroxypropyl-β-cyclodextrin (HP-β-CD) to enhance solubility and stability without chemical intervention.
F. The formation of these complexes was rigorously validated through UV–Vis, FT-IR, PXRD, TEM, and 2 H-NMR spectroscopy.
G. The evidence does not state that this conventional process yields sulfite, sulfate, and sulfide byproducts, leading to significant environmental and toxicological concerns.
H. This conventional process yields sulfite, sulfate, and sulfide byproducts, leading to significant environmental and toxicological concerns.


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
p -Coumaric acid (p-CA) is a key aromatic precursor for the biosynthesis of flavonoids, stilbenoids, and other high-value phenylpropanoids. While microbial production of p-CA typically relies on sugar-based substrates, methanol offers a sustainable and cost-effective alternative, though its use for aromatic biosynthesis remains unexplored. Here, we report the first de novo production of p-CA from methanol using engineered methylotrophic yeast Pichia pastoris . Through heterologous expression of a tyrosine ammonia-lyase and implementing a balanced push–pull strategy in the shikimate pathway using feedback-resistant variants of DAHP synthase ( ARO4 ) and chorismate mutase ( ARO7 ), carbon flux from methanol-derived C3 and C4 precursors was effectively redirected toward aromatic biosynthesis. Shake-flask studies revealed strong gene-dosage-dependent p-CA production, but strains with high-copy numbers suffered metabolic burden under high-density fermentation. Fed-batch bioreactor cultivation demonstrated that a moderate-copy strain achieved the highest titer of 704 ± 6 mg/L, outperforming high-copy variants in robustness and scalability. This study establishes P . pastoris as a promising chassis for methanol-based aromatic production and highlights the critical trade-off between pathway amplification and cellular fitness in C1 biomanufacturing. The online version contains supplementary material available at 10.1186/s40643-026-01068-7.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Tobacco waste, rich in nicotine, is both an environmental burden and a potential feedstock for high-value chemicals. Here, we developed a proximity-enhanced co-immobilized multi-enzyme cascade that efficiently converts the nicotine into a pharmaceutical intermediate 3-succinoylpyridine (SP). The cascade system comprises the nicotine oxidoreductase NicA2 V321 (NicAm), pseudooxynicotine amine oxidase (Pnao), and 3-succinoylsemialdehyde-pyridine dehydrogenase (Sapd) for sequential nicotine conversion, coupled with an aldehyde–ketone reductase (AKR) module for in situ NADP + regeneration. To improve nicotine-to-SP conversion and facilitate multi-enzyme recycling, a SpyCatcher/SpyTag self-assembled cofactor regeneration enzyme scaffold was adopted, in conjunction with AviTag–BirA-mediated biotinylation for cascade enzyme site-specific co-immobilization on streptavidin-coated supports. This proximity-enhanced design promoted efficient cofactor cycling and boosted the nicotine-to-SP conversion to approximately 63%, much higher than the 42.7% achieved by free enzymes. The co-immobilized system also showed improved pH and thermal stability, retaining over 60% of its initial activity after eight reuse cycles. This modular biocatalytic strategy provides a green and promising route for the valorization of nicotine-rich tobacco waste. The online version contains supplementary material available at 10.1186/s40643-026-01066-9.

Candidates:
A. Tobacco waste, rich in nicotine, is both an environmental burden and a potential feedstock for high-value chemicals.
B. Tobacco waste, rich in nicotine, is not both an environmental burden and a potential feedstock for high-value chemicals.
C. The cascade system comprises the nicotine oxidoreductase NicA3 V321 (NicAm), pseudooxynicotine amine oxidase (Pnao), and 3-succinoylsemialdehyde-pyridine dehydrogenase (Sapd) for sequential nicotine conversion, coupled with an aldehyde–ketone reductase (AKR) module for in situ NADP + regeneration.
D. To improve nicotine-to-SP conversion and facilitate multi-enzyme recycling, a SpyCatcher/SpyTag self-assembled cofactor regeneration enzyme scaffold was adopted, in conjunction with AviTag–BirA-mediated biotinylation for cascade enzyme site-specific co-immobilization on streptavidin-coated supports.
E. Here, we developed a proximity-enhanced co-immobilized multi-enzyme cascade that efficiently converts the nicotine into a pharmaceutical intermediate 3-succinoylpyridine (SP).
F. To improve nicotine-to-SP conversion and facilitate multi-enzyme recycling, a SpyCatcher/SpyTag self-assembled cofactor regeneration enzyme scaffold was not adopted, in conjunction with AviTag–BirA-mediated biotinylation for cascade enzyme site-specific co-immobilization on streptavidin-coated supports.
G. Here, we developed a proximity-enhanced co-immobilized multi-enzyme cascade that efficiently converts the nicotine into a pharmaceutical intermediate 4-succinoylpyridine (SP).
H. The cascade system comprises the nicotine oxidoreductase NicA2 V321 (NicAm), pseudooxynicotine amine oxidase (Pnao), and 3-succinoylsemialdehyde-pyridine dehydrogenase (Sapd) for sequential nicotine conversion, coupled with an aldehyde–ketone reductase (AKR) module for in situ NADP + regeneration.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["120 °C", "35%", "10 h", "36%", "2.75%", "11 h", "1.75%", "121 °C"]

Evidence:
This study assesses the impact of physicochemical pretreatment, enzymatic hydrolysis, and co-culture fermentation strategies on bioethanol production from corn husk biomass (CHB). Under optimal alkali pretreatment conditions (1.75% alkali, 4.0 g substrate concentration, 120 °C, 10 h), 35% lignin removal was achieved, with 48% cellulose and 39% hemicellulose recovery. In contrast, acid pretreatment resulted in 30% lignin removal, 45% cellulose recovery, and 34% hemicellulose recovery, showing lower efficiency than alkali pretreatment. During ultrasonication alkali pretreatment enhanced cellulose and hemicellulose exposure up to 51 and 46% and delignification up to 49%. Enzymatic hydrolysis of pretreated corn husk biomass was performed using commercial enzymes [Celluclast 1.5 L (700 EGU or 854 U mL −1 ) and Viscozyme (13.4 FBG/mL)] and isolated bacterial enzymes, including cellulase from Bacillus licheniformis (9.3 ± 0.3 U mL −1 ) and xylanase from Enterobacter asburiae PQ396173 (7.0 ± 0.4 U mL −1 ). The developed enzyme cocktail in ratio 3:2:3:1 (v/v; U mL −1 ) (Celluclast: Viscozyme: native cellulase: native xylanse) using a cocktail of native and commercial enzymes, yielded total reducing sugar of 740 mg g −1 glucose and 54.6 mg g −1 xylose. Fermentation of hydrolysate prepared with commercial enzymes using monoculture of Saccharomyces cerevisiae and Pichia pastoris yielded 17.6 g L −1 and 12.2 g L −1 bioethanol separately. Co-cultured yeasts produced 26.8 g L −1 ethanol at 96 h of incubation, exceeding monoculture yields. The fermentation with integration of commercial and isolated bacterial enzyme cocktails yielded the highest bioethanol output of 37.3 g L −1 at 96 h incubation, indicating that enzymatic saccharification with a combination of commercial and native enzyme cocktails results in maximum bioethanol production. The online version contains supplementary material available at 10.1186/s40643-026-01062-z. Keywords: Corn husk biomass, Ultrasonication pretreatment, Enzymatic hydrolysis, Enzyme cocktail, Fermentation, Bioethanol


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Brazzein is a heat-stable sweet protein with strong potential as a sugar alternative. Here, we leveraged precision fermentation to produce high titers of recombinant brazzein in the yeast Komagataella phaffii . As the brazzein expression cassette included a zeocin resistance marker, we investigated whether high zeocin resistance could serve as a predictive indicator of high brazzein production. A microscale screen across 100–1000 µg mL −1 zeocin rapidly identified clones with varying resistance, including the high-resistant Braz_3, which produced 4.5-fold more brazzein than the moderate-resistant Braz_7 in fed-batch bioreactors (61.5 vs. 13.7 mg L −1 ). Long-read genome sequencing revealed that five copies of the expression cassette were integrated into the genome of the high-producing strain compared to a single copy in the moderate-producing strain. Consequently, our results suggest a positive correlation between zeocin resistance and brazzein titers, demonstrating that antibiotic resistance phenotyping can guide strain selection for scalable production of functional food proteins. The online version contains supplementary material available at 10.1007/s10068-026-02150-8.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Increasing the production of structured cultivated meat (CM) remains a major challenge. Many current 3D printing and bioprinting technologies lack the necessary throughput, material compatibility, and cytocompatibility to produce realistic meat alternatives. In this study, screen printing, a traditional high-throughput printing technology, was investigated for the first time for the application of meat alternatives. The new process, 3D bio-screen printing (3D-BSP), presents a biomanufacturing process that uses edible materials (e.g., plant proteins) to produce meat-like structures with high resolution (0.1 mm). The suitability of edible inks for the 3D-BSP process was investigated on the basis of soy protein isolate (SPI) rheologically (flow index value < 0.4) and in terms of printability. In order to increase the protein content in edible inks while maintaining processability, an approach based on reducing agents was investigated. Sodium sulphite served as a demonstration model and resulted in a high protein content (>20 wt%) in SPI, while maintaining the flow properties suitable for processing protein-rich inks. To test the suitability as scaffolding technology for hybrid cultivated meat, meat-like scaffold structures were printed with C2C12 myoblasts differentiated on them. The printability of the structures was high in the resolution range investigated from 0.1 mm to 1 mm and supported 2D and 3D myoblast cultures (64% actin coverage and myotube formation). Ultimately, a marbled prototype was produced whose thickness could be increased by a stacking approach (>0.5cm). The texture profile (e.g., chewiness) of stacked and printed scaffolds was comparable to that of conventional meat. Upon successful transfer of the process parameters determined here to industrial screen printing machines, production rates of >100 kg/h could be achieved with one machine in the future. 3D-BSP offers a practical and cost-effective perspective for the mass production of structured meat in industrial quantities using screen printing technology. It could remove technical and economic barriers in this area and bring us closer to the commercial production of high-quality meat alternatives.

Candidates:
A. The evidence does not state that increasing the production of structured cultivated meat (CM) remains a major challenge.
B. In this study, screen printing, a traditional high-throughput printing technology, was not investigated for the first time for the application of meat alternatives.
C. The new process, 4D bio-screen printing (3D-BSP), presents a biomanufacturing process that uses edible materials (e.g., plant proteins) to produce meat-like structures with high resolution (0.1 mm).
D. The new process, 3D bio-screen printing (3D-BSP), presents a biomanufacturing process that uses edible materials (e.g., plant proteins) to produce meat-like structures with high resolution (0.1 mm).
E. Increasing the production of structured cultivated meat (CM) remains a major challenge.
F. Many current 3D printing and bioprinting technologies lack the necessary throughput, material compatibility, and cytocompatibility to produce realistic meat alternatives.
G. In this study, screen printing, a traditional high-throughput printing technology, was investigated for the first time for the application of meat alternatives.
H. Many current 4D printing and bioprinting technologies lack the necessary throughput, material compatibility, and cytocompatibility to produce realistic meat alternatives.


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Human induced pluripotent stem cells (iPSCs) are transforming adoptive cell therapy by combining unlimited self-renewal, broad differentiation potential, and high amenability to genome engineering. These attributes make iPSCs a versatile source for the development of standardized immune effector cells at industrial scale, enabling a shift from patient- or donor-restricted cell products toward true off-the-shelf immunotherapies that can be improved through iterative genome engineering. iPSC-derived natural killer (iNK) cells are the most clinically advanced and exemplify the platform’s advantages over conventional autologous or donor-sourced approaches. Unlike autologous therapies, which require labor-intensive and expensive personalized clinical-grade manufacturing, and are constrained by variable quality and genetic intractability of donor products, iPSC technology supports the creation of renewable, clonally defined master cell banks as uniform starting material for NK-cell therapy products. Advances in CRISPR/Cas-based editing now permit multiplex introduction of functional traits, enhanced cytokine signaling, antibody-dependent cytotoxicity, checkpoint resistance, optimized trafficking, safety switches, and increasing signal complexity, directly at the pluripotent or progenitor stages; ultimately allowing for fully-programmable iNK cells with customizable potency and persistence. Early clinical studies of iNK products validate the feasibility, safety, and therapeutic potential of this approach, but also underscore the need for continued refinement of differentiation protocols, manufacturing pipelines, and regulatory standards to ensure efficacy, genomic stability, phenotypic maturity, and long-term safety. This review outlines current breakthroughs and future directions of iNK cell therapies, emphasizing how programmable iPSC chassis platforms are enabling modular and off-the-shelf targeted immunotherapies.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["30 °C", "31 °C", "5.4%", "4.4%"]

Evidence:
To achieve high-value utilization of brewer’s spent grains and to produce protein-rich food ingredients, brewer’s spent grains were upcycled by a liquid fungal fermentation and the fermentation conditions were optimized. Brewer’s spent grains (BSG) represent the most abundant by-product of the brewing industry. Although BSG are food-grade, their direct use in food is limited because of sensory changes of the products. In this study, screening experiments revealed that black beer spent grains could be efficiently upcycled by submerged fermentation with the edible fungus Pleurotus ostreatus . The conditions of the fermentation of BSG with P. ostreatus were optimized using response surface methodology, including the parameters substrate concentration, inoculum volume, initial pH value, and temperature. As no separation between BSG and mycelium was possible after the fermentation, ergosterol was used as a biomarker to determine the fungal growth. As optimum conditions, a BSG concentration of 17 g L − 1 dry matter, an inoculum volume of 4.4% (v/v), an initial pH of 9.3 and a temperature of 30 °C were identified. The fermentation with P. ostreatus increased the true protein content of the biomass compared to the spent grains and reduced the total fat content. The biological value was increased from 88 to 94 (reference standard is whole chicken egg with a biological value of 100). Tryptophan and lysine were limiting in the non-fermented spent grains, while after fermentation, the chemical score of lysine increased from 78 to 92, and tryptophan wasn’t limiting anymore. Fermentation by P. ostreatus enhanced the nutritional value of BSG, and the fermentation conditions were optimized by using response surface methodology. The online version contains supplementary material available at 10.1186/s40643-026-01018-3.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["98%", "94%", "99%", "93%"]

Evidence:
Ultrafiltration and diafiltration (UF/DF) operations have been demonstrated to clear leachables from drug substance, however there is limited data available. Consequently, comprehensive and systematic characterization of leachables clearance during UF/DF is required and essential. To achieve this, the reduction capacity for 28 selected organic compounds spiked into 3 different proteins during UF/DF processes was investigated using liquid chromatography high-resolution mass spectrometry. Selection of compounds was based on their presence in representative biomanufacturing processes. Most compounds (24) showed clearance over 98% across the process for the 3 protein materials. The specific protein characteristics and process parameters for each protein had a minimal impact on clearance, with sieving coefficients essentially the same for each one of the 3 protein processes. The sieving coefficient is a parameter that characterizes clearance of compounds during UF/DF. Physicochemical properties of the compounds under study significantly influenced their clearance, with the octanol–water coefficient (Log P) being the most crucial factor. Compounds with Log P < 4 had sieving coefficients close to ideal clearance, and compounds with Log P > 7 showed lower but still significant clearance (> 93%). Other important parameters were established to be molecular weight, polarizability and solvent accessible surface area. Modelling tools based on Orthogonal Partial Least Squares (OPLS) regression were created to predict sieving coefficients. The present work has created a strong background to describe the ability of UF/DF to remove potential organic leachables. Application of these modelling approaches becomes critical to support product safety assessments. Demonstration of significant removal along UF/DF operations confirms risk reduction of leachables coming mostly from upstream stages. The online version contains supplementary material available at 10.1007/s11095-026-04050-2. Keywords: Liquid chromatography-mass spectrometry, Process equipment related leachables, Regression models, Ultrafiltration/diafiltration, Vortex-assisted liquid–liquid microextraction


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Polyethylene terephthalate (PET) plastic waste causes serious environmental pollution due to insufficient recycling rates. Enzymatic PET depolymerization offers a sustainable recycling strategy, but limited stability and activity of current PET-degrading enzymes restrict practical implementation. Here, we engineer Polyester Hydrolase Leipzig 7 (PHL7), a PET hydrolase from a compost metagenome, to enhance its stability and catalytic performance under recycling-relevant conditions. Using Rosetta PROSS-based computational design combined with rational mutagenesis, we introduce up to 24 mutations, generating variants with melting temperatures of 88-95 °C and over 110-fold higher activity in 0.1 M phosphate buffer compared to the parent enzyme. Benchmarking shows that the best variants (R4M6, R4M9, and R4M10) match or exceed the performance of established engineered PET hydrolases, including ICCG and LCC-A2, and approach that of TurboPETase across multiple conditions. Under high substrate loadings, the PHL7-R4 variants degrade 75-78% of 10% (w/w) PET within 24 h at 65 °C, outperforming ICCG, while an optimized variant R4M10-H185Y achieves up to 84% degradation of 20% (w/w) PET. X-ray structure determination and molecular dynamics simulations reveal key stabilizing and activity enhancing mechanisms. These engineered PHL7 variants represent robust biocatalysts for scalable enzymatic PET recycling.

Candidates:
A. Here, we engineer Polyester Hydrolase Leipzig 8 (PHL7), a PET hydrolase from a compost metagenome, to enhance its stability and catalytic performance under recycling-relevant conditions.
B. Polyethylene terephthalate (PET) plastic waste causes serious environmental pollution due to insufficient recycling rates.
C. The evidence does not state that polyethylene terephthalate (PET) plastic waste causes serious environmental pollution due to insufficient recycling rates.
D. Enzymatic PET depolymerization offers a sustainable recycling strategy, but limited stability and activity of current PET-degrading enzymes restrict practical implementation.
E. Using Rosetta PROSS-based computational design combined with rational mutagenesis, we introduce up to 25 mutations, generating variants with melting temperatures of 88-95 °C and over 110-fold higher activity in 0.1 M phosphate buffer compared to the parent enzyme.
F. Here, we engineer Polyester Hydrolase Leipzig 7 (PHL7), a PET hydrolase from a compost metagenome, to enhance its stability and catalytic performance under recycling-relevant conditions.
G. Using Rosetta PROSS-based computational design combined with rational mutagenesis, we introduce up to 24 mutations, generating variants with melting temperatures of 88-95 °C and over 110-fold higher activity in 0.1 M phosphate buffer compared to the parent enzyme.
H. The evidence does not state that enzymatic PET depolymerization offers a sustainable recycling strategy, but limited stability and activity of current PET-degrading enzymes restrict practical implementation.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["11%", "51%", "50%", "10%"]

Evidence:
When applied for industrial-scale bioproduction, cells are subjected to ever-changing cultivation environments due to bioreactor heterogeneities, which can significantly influence their growth and production behavior. Cultivating and comparing cells and strains under conditions representative of an actual bioprocess instead of laboratory environments is therefore necessary for developing more reliable production strains. Scale-down bioreactors provide an experimental approach for this endeavor, yet they are limited in their temporal resolution and flexibility when it comes to environmental dynamics. Thus, the impact of second-scale differences in dynamic environments on microbial producers remains unexplored. This study uses the advantages of microfluidic single-cell cultivation to compare the growth behavior and intracellular parameters of three yeast strains under dynamic cultivation conditions with alternating phases of glucose excess and limitation at the timescale of seconds aiming to uncover strain-specific performance under as well as adaptation to fluctuating, bioprocess-relevant environments. Across all three strains, a consistent trend was observed: decreasing glucose availability resulted in reduced growth rates, lower ATP levels, diminished glycolytic flux, and smaller cell sizes. Notably, cells already reached approximately 50% of their maximal growth rate when exposed to growth-promoting glucose conditions for only 10% of the time. Importantly, both growth rate and cell size exhibited an adaptation phase rather than an immediate response to oscillatory glucose supply. Longer exposure to favorable conditions shortened the adaptation phase and resulted in higher adapted growth rates and larger cell sizes. By leveraging microfluidic single-cell cultivation, this study provides unprecedented temporal resolution of environmental dynamics, enabling direct, strain-specific comparison of cellular responses and adaptation to rapidly changing cultivation environments. Seconds do make a difference. Our findings demonstrate that growth rate is a highly conserved trait under environmental perturbations, which points to an intrinsic robustness of the investigated strains and highlights adaptation as a key determinant of performance in dynamic environments. Together, these insights have important implications for the design of future single-cell experiments and for the development of robust bioprocesses operating under dynamic conditions. The online version contains supplementary material available at 10.1186/s12934-026-03029-3. Keywords: Dynamic microfluidic single-cell cultivation, Strain comparison, Bioprocess development, Saccharomyces cerevisiae, Biosensors, Scale-down, Dynamic environment


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Carboxylic acids are key platform chemicals that can be biologically produced in mixed-culture fermentations. In these systems, product formation is determined by the microbial community composition, which is shaped by operational conditions. A general trend has been observed linking substrate availability to the production of either lactic acid or volatile fatty acids, but it remains unclear at which substrate concentration this shift occurs. This study investigates the effect of carbohydrate concentrations (from 0 to 670 mg COD·L −1 ) and type (hexoses and pentoses) on product selectivity and microbial community composition in thermophilic mixed-culture fermentations. Fermentation experiments were conducted in thermophilic reactors (55 °C, pH 5.3, HRT 4 days), fed either with glucose or xylose. Higher substrate concentrations (650–670 mg COD·L −1 ) favored lactic acid production, accounting for 60–76% of the soluble products. Lower carbohydrate concentrations reduced lactic acid production and increased volatile fatty acid concentrations. Volatile fatty acids became the main product under low substrate concentrations (continuous operation; 0 mg COD·L −1 ), with a selectivity of 83–97%. Despite this general trend, the substrate concentration at which the shift from lactic acid to volatile fatty acids occurred depended on the carbohydrate type. Microbial community analyses revealed Thermoanaerobacterium as the dominant genus in all reactors, with genera within the Bacillaceae family (putatively involved in lactic acid production) increasing in relative abundance under high carbohydrate concentrations. This suggests that the product shift resulted from both a metabolic shift within the dominant species and a change in the microbial community composition. Low carbohydrate concentrations favor VFA production. High carbohydrate concentrations promote lactic acid formation. Carbohydrate type influences the shift from lactic acid to VFAs. Low carbohydrate concentrations favor VFA production. High carbohydrate concentrations promote lactic acid formation. Carbohydrate type influences the shift from lactic acid to VFAs. The online version contains supplementary material available at 10.1007/s00253-026-13806-0.


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Surfactin production by Bacillus subtilis is typically performed under aerobic conditions, requiring high aeration and agitation, which leads to mechanical stress for the cells and therefore promotes excessive foaming. As a process-oriented alternative to full aerobic operation, an aerobic to micro-aerobic switching strategy was developed that aims to decouple biomass formation from surfactin synthesis by controlling oxygen availability. Promoter activities relevant to nitrate respiration and surfactin biosynthesis were first characterised in shake flasks using transcriptional reporter strains and online monitoring of dissolved oxygen. The nitrite reductase promoter showed the strongest induction under oxygen-limited conditions and was used to construct an oxygen-responsive production strain in which the native P srfA promoter was replaced to suppress surfactin formation during aerobic growth and shift production to the micro-aerobic phase. The switching concept was subsequently evaluated in 30-L stirred-tank bioreactor cultivations using stepwise reductions of dissolved oxygen setpoints combined with exponential glucose feeding and nitrate supplementation. The engineered strain B. subtilis MG19 enabled stable transitions into micro-aerobic operation without apparent loss of biomass, while a reference strain B. subtilis MG17 with the native regulation showed surfactin formation already during the aerobic phase and pronounced process disturbance after switching, characterised by glucose accumulation and a strong decrease in surfactin concentration. Overall, the study demonstrates oxygen switching as a scalable process engineering tool for controlling surfactin production phases and highlights key constraints for robust micro-aerobic bioreactor operation. • Oxygen availability decoupled growth and surfactin formation. • Micro-aerobic process control was evaluated in 30-L bioreactor cultivations. • The engineered strain shifted surfactin production to oxygen-limited conditions. The online version contains supplementary material available at 10.1007/s00253-026-13868-0. Keywords: Lipopeptide production, Oxygen-limited cultivation, Fed-batch fermentation, Bioreactor scale-up, Process kinetics, Nitrogen metabolism


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
As the technological advancements of the early 21st century are pushing industrial biotechnology (IB) into the realm of Big Data–driven innovation, the requirement for trustworthy data management, annotation, and standardization is emerging as a necessity. Minimum information models (MIMs) have long been used across disciplines as the backbone of good data management practices by providing the scaffold upon which standardized recording of metadata can adequately and succinctly describe an understudied phenomenon. Here we present a minimum set of metadata, named the minimum information for fermentation experiments (MIFE) and devices (MIFD), that has been specifically designed to accommodate the data management and annotation needs of IB-related fermentation experiments. Although the proposed schema is tailored to IB applications, MIFE and MIFD build upon well-established models and community standards to facilitate easier integration with existing infrastructure and easier adoption by the community, as well as aim to integrate Findable, Accessible, Interoperable, and Reproducible (FAIR) principles in the IB field. In addition, the integration with FAIR Data Station (FAIR DS), a tool that offers metadata validation and enables the automated uptake of (meta)data from data management repositories such as FAIRDOM-SEEK, is showcased. The proposed models are accompanied by a Python package that enables their programmatic use by creating a Linked Data Modeling Language (LinkML) schema that can fuel subsequent analyses. Through the promotion and simplification of knowledge discovery, we believe that MIFE and MIFD can accelerate the application of state-of-the-art artificial intelligence (AI) methods and the adoption of explainable AI to better understand bioprocesses at scale.

Candidates:
A. Minimum information models (MIMs) have long been used across disciplines as the backbone of good data management practices by providing the scaffold upon which standardized recording of metadata cannot adequately and succinctly describe an understudied phenomenon.
B. The evidence does not state that here we present a minimum set of metadata, named the minimum information for fermentation experiments (MIFE) and devices (MIFD), that has been specifically designed to accommodate the data management and annotation needs of IB-related fermentation experiments.
C. Although the proposed schema is not tailored to IB applications, MIFE and MIFD build upon well-established models and community standards to facilitate easier integration with existing infrastructure and easier adoption by the community, as well as aim to integrate Findable, Accessible, Interoperable, and Reproducible (FAIR) principles in the IB field.
D. As the technological advancements of the early 21st century are pushing industrial biotechnology (IB) into the realm of Big Data–driven innovation, the requirement for trustworthy data management, annotation, and standardization is emerging as a necessity.
E. Although the proposed schema is tailored to IB applications, MIFE and MIFD build upon well-established models and community standards to facilitate easier integration with existing infrastructure and easier adoption by the community, as well as aim to integrate Findable, Accessible, Interoperable, and Reproducible (FAIR) principles in the IB field.
F. Minimum information models (MIMs) have long been used across disciplines as the backbone of good data management practices by providing the scaffold upon which standardized recording of metadata can adequately and succinctly describe an understudied phenomenon.
G. As the technological advancements of the early 22st century are pushing industrial biotechnology (IB) into the realm of Big Data–driven innovation, the requirement for trustworthy data management, annotation, and standardization is emerging as a necessity.
H. Here we present a minimum set of metadata, named the minimum information for fermentation experiments (MIFE) and devices (MIFD), that has been specifically designed to accommodate the data management and annotation needs of IB-related fermentation experiments.


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["50 °C", "77 h", "1386 h", "51 °C", "76 h", "1387 h"]

Evidence:
Carbocyclic nicotinamide cofactors are attractive NAD(P) + analogues that retain native redox activity while offering substantially enhanced chemical and thermal stability. Their broader use, however, has been limited by demanding multistep chemical syntheses involving complex protection strategies and limited exploration of enzymatic alternatives. In this study, we studied both chemical and enzymatic approaches for the synthesis of carba-analogues. For the enzymatic route, we identify and characterize the key enzymes involved in cofactor assembly, evaluating their tolerance toward non-native, carbocyclic substrates and extending the assembly to further generate the phosphorylated analogue, cNADP + . In addition, extending the analysis to cofactor thermostability, carba-NAD + displayed a remarkable halftime ( t 1/2 > 1386 h at 50 °C), far exceeding that of NAD + ( t 1/2 = 76 h) and is accepted by a broad panel of oxidoreductases. Collectively, this work outlines a modular workflow and details the synthesis landscape for accessing thermostable, synthetic nicotinamide cofactor analogues, cNAD + and cNADP + , unveiling new opportunities for their application in cofactor engineering and synthetic biocatalysis.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Geological carbon sequestration (GCS) is a key option for climate change mitigation, and subsurface microorganisms can alter CO 2 behavior through biomethanation, bioliquefaction, and biomineralization. This review summarizes the major microbial processes involved in GCS and evaluates their effects on carbon stabilization, resource reutilization, and storage risk. We compare microbial distribution and metabolic functions across representative geological reservoirs and synthesize laboratory, numerical, and field approaches into a multi-scale framework for studying microbially mediated GCS. We also discuss engineering regulation strategies, site selection, monitoring, and risk control, together with current technical challenges and future research priorities. Overall, this review provides an integrated perspective on microbial mechanisms and practical guidance for safer and more effective GCS deployment. Subject areas: Earth sciences, Biogeochemistry, Biogeoscience, Global carbon cycle, Microbiology, Applied microbiology

Candidates:
A. Geological carbon sequestration (GCS) is a key option for climate change mitigation, and subsurface microorganisms can alter CO 2 behavior through biomethanation, bioliquefaction, and biomineralization.
B. Geological carbon sequestration (GCS) is a key option for climate change mitigation, and subsurface microorganisms can alter CO 3 behavior through biomethanation, bioliquefaction, and biomineralization.
C. The evidence does not state that this review summarizes the major microbial processes involved in GCS and evaluates their effects on carbon stabilization, resource reutilization, and storage risk.
D. We compare microbial distribution and metabolic functions across representative geological reservoirs and synthesize laboratory, numerical, and field approaches into a multi-scale framework for studying microbially mediated GCS.
E. The evidence does not state that we compare microbial distribution and metabolic functions across representative geological reservoirs and synthesize laboratory, numerical, and field approaches into a multi-scale framework for studying microbially mediated GCS.
F. We also discuss engineering regulation strategies, site selection, monitoring, and risk control, together with current technical challenges and future research priorities.
G. The evidence does not state that we also discuss engineering regulation strategies, site selection, monitoring, and risk control, together with current technical challenges and future research priorities.
H. This review summarizes the major microbial processes involved in GCS and evaluates their effects on carbon stabilization, resource reutilization, and storage risk.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The development of microbial biocomposites represents a promising frontier in the design of sustainable, multifunctional, and environmentally friendly materials. This minireview synthesises recent advances (2020–2026) in the production, modification, and application of biocomposites derived from microorganisms, with a focus on bacterial cellulose (BC) and fungal mycelium. It explores their synergies with emerging approaches in synthetic biology, 3D printing, and mineral functionalisation. Key microbial systems such as Komagataeibacter spp. and Ganoderma spp. are discussed, alongside structural and functional engineering strategies including in situ hydroxyapatite mineralisation, incorporation of plant fibres, and the addition of functional nanomaterials such as graphene oxide. The review further highlights the integration of these materials into high-value applications, including osteogenic scaffolds, self-healing living materials, biodegradable packaging, and environmental remediation systems. Finally, it addresses current regulatory and technical challenges related to industrial scalability, functional stability, and batch-to-batch standardisation. This article aims to provide a critical and comprehensive perspective for researchers and professionals in applied microbiology, materials science, and industrial biotechnology, emphasising the potential of microbial biocomposites as a convergent platform at the interface of sustainability, functional innovation, and bioinspired design.

Candidates:
A. It explores their synergies with emerging approaches in synthetic biology, 4D printing, and mineral functionalisation.
B. It explores their synergies with emerging approaches in synthetic biology, 3D printing, and mineral functionalisation.
C. The development of microbial biocomposites represents a promising frontier in the design of sustainable, multifunctional, and environmentally friendly materials.
D. Key microbial systems such as Komagataeibacter spp.
E. This minireview synthesises recent advances (2020–2026) in the production, modification, and application of biocomposites derived from microorganisms, with a focus on bacterial cellulose (BC) and fungal mycelium.
F. The evidence does not state that the development of microbial biocomposites represents a promising frontier in the design of sustainable, multifunctional, and environmentally friendly materials.
G. The evidence does not state that key microbial systems such as Komagataeibacter spp.
H. This minireview synthesises recent advances (2021–2026) in the production, modification, and application of biocomposites derived from microorganisms, with a focus on bacterial cellulose (BC) and fungal mycelium.


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Aspergillus welwitschiae strain SVUAw9 was isolated from sugarcane bagasse and identified by the sequencing of the calmodulin (CaM) gene, was found to exhibit substantial cold-active xylanase activity in this study. Under solid-state fermentation conditions, the strain can utilize a range of agro-industrial residues as substrates to produce cold-active xylanase. After 14 days at pH 7 and 10 °C using ammonium chloride as the nitrogen source, sugarcane bagasse was the most productive substrate, generating 106.93 ± 12.1 U/gram dry substrate (gds), followed by bean straw, corn cob, and date palm leaves which produced 57.82 ± 8.27, 57.75 ± 7.44, and 38.87 ± 6.15 U/gds, respectively. Conversely, the substrate exhibiting the lowest production was rice husk, producing 29.32 ± 4.88 U/gds. Using Trilite MC 08 and Sephacryl S-200 columns, the xylanase was purified 75.87 times, yielding a single band at approximately 71 kDa. At pH 4.0 and 30 °C, the maximum activity of 156.46 ± 12 U/mg was achieved. Co(NO₃)₂, MnSO₄, and NiSO₄ markedly enhanced enzyme activity, resulting in residual activity increases of 152.94 ± 11.54%, 152.94 ± 9.58%, and 134.12 ± ठर 7.66%, respectively, with specific activities of 239.3 ± 18, 239.3 ± 15, and 209.84 ± 12 U/mg. Km and Vmax for the pure xylanase were determined as 0.1 ± 0.005 mg/mL and 144.9 ± 7.14 µmol/min, respectively. The purified xylanase could degrade oat spelt xylan, corn cob xylan, xylan, Birchwood xylan, maize stalk xylan, carboxymethyl cellulose (CMC), beechwood and microcrystalline cellulose (MCC), resulting in activities of 156 ± 12, 108 ± 7, 82 ± 6, 79 ± 5, 62 ± 4, 22 ± 1.5, and 38 ± 3 U/mg, respectively. Keywords: Cold-adapted, Bagasse , Enzymes, Fungi, Lignocellulose, Plant-biomass-derived waste, Xylanase


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The chemical and biochemical engineering, microbiology, and biotechnology communities deeply mourn the passing of Dr. Robert M. Kelly, the Alcoa Professor of Chemical and Biomolecular Engineering and Director of the Biotechnology Program at NC State University. A towering pioneer in the biology of extremophiles, an esteemed educator and mentor, and a dedicated editor for Applied and Environmental Microbiology (AEM), Bob leaves behind an extraordinary legacy that reshaped our understanding of life at high temperatures.

Candidates:
A. The chemical and biochemical engineering, microbiology, and biotechnology communities deeply mourn the passing of Dr.
B. The evidence does not state that kelly, the Alcoa Professor of Chemical and Biomolecular Engineering and Director of the Biotechnology Program at NC State University.
C. A towering pioneer in the biology of extremophiles, an esteemed educator and mentor, and a dedicated editor for Applied and Environmental Microbiology (AEM), Bob leaves behind an extraordinary legacy that reshaped our understanding of life at high temperatures.
D. The evidence does not state that a towering pioneer in the biology of extremophiles, an esteemed educator and mentor, and a dedicated editor for Applied and Environmental Microbiology (AEM), Bob leaves behind an extraordinary legacy that reshaped our understanding of life at high temperatures.
E. Kelly, the Alcoa Professor of Chemical and Biomolecular Engineering and Director of the Biotechnology Program at NC State University.
F. The evidence does not state that the chemical and biochemical engineering, microbiology, and biotechnology communities deeply mourn the passing of Dr.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Industrial production of malic acid remains dependent on fossil resources or, when performed microbiologically, on sugar-based feedstocks. Both routes come with caveats, generating emissions and competing with food supply. The use of CO₂-derived one-carbon substrates offers a promising alternative to circumvent these constraints. In this study, the malic acid production process from methanol in metabolically engineered Ogataea polymorpha NYCY495 LEU-ΔSTE12 Pyc Mdh MAE1 strain was optimized and scaled up. A two-phase cultivation strategy, using glycerol for biomass formation and methanol for product synthesis, was established in shake flasks and subsequently transferred to a 1 L bioreactor. Process optimization through automated feeding strategies was evaluated. DO-based feeding was the most effective approach, using a combination of methanol and glycerol, achieving a final molar yield of 0.1 mol MA mol MeOH ⁻¹ and a maximum productivity of 0.5 g l⁻¹ h⁻¹. This successful fermentation strategy was validated using green methanol, showcasing the feasibility of “closing the loop” as envisioned in the bioeconomy. Finally, a comparative study of the effect of glycerol, methanol, and their mixture on O. polymorpha NYCY495 LEU-ΔSTE12 Pyc Mdh MAE1 methanol metabolism, peroxisome biogenesis, and cellular redox balance is presented, supporting the positive cumulative effect of both on gene transcription. Keywords: C1 metabolism, methylotrophic yeast, bioprocess optimization, cofeeding, green methanol, transcriptomics

Candidates:
A. The use of CO₂-derived one-carbon substrates offers a promising alternative to circumvent these constraints.
B. Industrial production of malic acid remains dependent on fossil resources or, when performed microbiologically, on sugar-based feedstocks.
C. The evidence does not state that the use of CO₂-derived one-carbon substrates offers a promising alternative to circumvent these constraints.
D. The evidence does not state that both routes come with caveats, generating emissions and competing with food supply.
E. In this study, the malic acid production process from methanol in metabolically engineered Ogataea polymorpha NYCY495 LEU-ΔSTE12 Pyc Mdh MAE1 strain was optimized and scaled up.
F. The evidence does not state that industrial production of malic acid remains dependent on fossil resources or, when performed microbiologically, on sugar-based feedstocks.
G. In this study, the malic acid production process from methanol in metabolically engineered Ogataea polymorpha NYCY496 LEU-ΔSTE12 Pyc Mdh MAE1 strain was optimized and scaled up.
H. Both routes come with caveats, generating emissions and competing with food supply.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Formaldehyde-based fixatives, particularly neutral-buffered formalin (NBF), are widely used in histology owing to their strong protein cross-linking capacity and reliable preservation of cytoarchitecture. However, formaldehyde is toxic and carcinogenic, motivating the search for safer alternatives. Here, we systematically evaluated lactic acid (LA) as a potential formaldehyde-free fixative for murine brain tissues. Brains were fixed either by immersion or by transcardial perfusion followed by immersion using LA at varying concentrations, pH-adjusted LA, or phosphate-buffered saline (PBS), and were compared with NBF. Histomorphological preservation was assessed using qualitative evaluation, semi-quantitative scoring, and quantitative crack-area measurements. Across all conditions, NBF provided the best overall tissue preservation. LA fixation showed clear concentration-dependent effects: low concentrations resulted in poor structural integrity, whereas higher concentrations (LA10–LA20) achieved moderate preservation of cytoarchitecture, reduced cracking, and acceptable cellular morphology. White-matter integrity remained inferior to NBF, consistent with the lipid-rich composition of myelin and the predominantly protein-denaturing, non-cross-linking fixation properties of LA. pH-adjustment experiments indicated that fixation efficacy partially depended on acidity. Notably, LA20 delivered by transcardial perfusion substantially improved tissue preservation compared with immersion alone and outperformed PBS. Although LA cannot fully replace NBF, high-concentration LA might represent an alternative for selected neurohistological applications. The online version contains supplementary material available at 10.1038/s41598-026-51513-y. Keywords: Histology, Fixation, Mouse brain, Neutral buffered formaldehyde, Paraffin sections, Tissue preservation Subject terms: Biological techniques, Medical research, Neurology, Neuroscience

Candidates:
A. The evidence does not state that here, we systematically evaluated lactic acid (LA) as a potential formaldehyde-free fixative for murine brain tissues.
B. Brains were not fixed either by immersion or by transcardial perfusion followed by immersion using LA at varying concentrations, pH-adjusted LA, or phosphate-buffered saline (PBS), and were compared with NBF.
C. Here, we systematically evaluated lactic acid (LA) as a potential formaldehyde-free fixative for murine brain tissues.
D. Brains were fixed either by immersion or by transcardial perfusion followed by immersion using LA at varying concentrations, pH-adjusted LA, or phosphate-buffered saline (PBS), and were compared with NBF.
E. Formaldehyde-based fixatives, particularly neutral-buffered formalin (NBF), are not widely used in histology owing to their strong protein cross-linking capacity and reliable preservation of cytoarchitecture.
F. However, formaldehyde is toxic and carcinogenic, motivating the search for safer alternatives.
G. However, formaldehyde is not toxic and carcinogenic, motivating the search for safer alternatives.
H. Formaldehyde-based fixatives, particularly neutral-buffered formalin (NBF), are widely used in histology owing to their strong protein cross-linking capacity and reliable preservation of cytoarchitecture.


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
The plant endoplasmic reticulum (ER) is a dynamic organelle composed of multiple distinct structural domains, such as cisternae, which are maintained by ER morphogens including the Arabidopsis thaliana Lunapark proteins (LNPs). Cisternae are typically described as sac‐like structures connected by tubules. Here we challenge this assumption and propose that cisternae have a more complex structure that modifies ER functionality. This study used state‐of‐the‐art high‐resolution confocal and variable‐angle epifluorescence microscopy, along with transmission electron microscopy and tomography, on high‐pressure frozen Arabidopsis thaliana samples. We found that AtLNP1‐stabilised ER forms cisternae composed of dense tubular matrices, whereas AtLNP2 forms cisternae with a uniform, sac‐like structure. Furthermore, overexpression of AtLNP proteins alters Golgi morphology, affecting ER‐to‐Golgi transport and secretion. Our findings reveal that the balance between AtLNP1 and AtLNP2 is critical for ER cisternae organisation and ER functionality in protein production and secretion. This work provides new insights into ER structural plasticity and its functional implications in plant cells. The plant endoplasmic reticulum (ER) is a dynamic organelle composed of multiple distinct structural domains, such as cisternae, which are maintained by ER morphogens including the Arabidopsis thaliana Lunapark proteins (LNPs). Cisternae are typically described as sac‐like structures connected by tubules. Here we challenge this assumption and propose that cisternae have a more complex structure that modifies ER functionality. This study used state‐of‐the‐art high‐resolution confocal and variable‐angle epifluorescence microscopy, along with transmission electron microscopy and tomography, on high‐pressure frozen Arabidopsis thaliana samples. We found that AtLNP1‐stabilised ER forms cisternae composed of dense tubular matrices, whereas AtLNP2 forms cisternae with a uniform, sac‐like structure. Furthermore, overexpression of AtLNP proteins alters Golgi morphology, affecting ER‐to‐Golgi transport and secretion. Our findings reveal that the balance between AtLNP1 and AtLNP2 is critical for ER cisternae organisation and ER functionality in protein production and secretion. This work provides new insights into ER structural plasticity and its functional implications in plant cells.

Candidates:
A. The evidence does not state that here we challenge this assumption and propose that cisternae have a more complex structure that modifies ER functionality.
B. The evidence does not state that this study used state‐of‐the‐art high‐resolution confocal and variable‐angle epifluorescence microscopy, along with transmission electron microscopy and tomography, on high‐pressure frozen Arabidopsis thaliana samples.
C. Here we challenge this assumption and propose that cisternae have a more complex structure that modifies ER functionality.
D. This study used state‐of‐the‐art high‐resolution confocal and variable‐angle epifluorescence microscopy, along with transmission electron microscopy and tomography, on high‐pressure frozen Arabidopsis thaliana samples.
E. The plant endoplasmic reticulum (ER) is not a dynamic organelle composed of multiple distinct structural domains, such as cisternae, which are maintained by ER morphogens including the Arabidopsis thaliana Lunapark proteins (LNPs).
F. Cisternae are typically described as sac‐like structures connected by tubules.
G. The plant endoplasmic reticulum (ER) is a dynamic organelle composed of multiple distinct structural domains, such as cisternae, which are maintained by ER morphogens including the Arabidopsis thaliana Lunapark proteins (LNPs).
H. Cisternae are not typically described as sac‐like structures connected by tubules.


Prompt:
Classify every fixed-vocabulary term as present or absent in the evidence. Vocabulary: bioreactor, cell culture, fermentation, chromatography, purification, filtration, viability, titer, potency, sterility, viral vector, gene therapy, cell therapy, monoclonal antibody, recombinant protein, dissolved oxygen. Return JSON with exactly two alphabetized keys: {"absent":[...],"present":[...]}. Preserve vocabulary order within each list. JSON whitespace is ignored.

Evidence:
Biocalorimetry offers a powerful approach for real-time process monitoring and optimization in the biotechnological utilization of both natural and synthetic macromolecules, particularly in complex solid-state systems aiming at the valorization of plant biomass or plastics’ waste. This review critically examines general strengths and limitations of biothermodynamics and calorimetry for monitoring microbial activity, which can be tracked across diverse scales and substrates via metabolic heat measurements. Metabolic heat-derived activity parameters enable the robust quantitative evaluation of the performance of microorganisms for substrate conversion, hence potentially representing valuable tools for bioprocess development and operation. While biocalorimetry is established in liquid-phase cultivation systems to some extent, its adaptation to solid-state fermentation, composting, and the biochemical breakdown of solid plastics still remains in early stages while holding promise for real-time control and upscaling. Challenges and limits of the applicability of this technology currently persist especially for mixed cultures and non-sterile processes. Nevertheless, expanding metabolic heat-based datasets to microbial functional traits could advance ecological and industrial applications. Overall, biocalorimetry is positioned as a valuable tool for advancing circular bioeconomy strategies, though further validation and methodological development are needed for broader adoption in both research and industrial contexts. • Biocalorimetry reliably quantifies microbial activity on complex solid substrates . • Biocalorimetry can support plant biomass and plastic waste valorization in a circular bioeconomy . • Biocalorimetric monitoring is applicable from the laboratory to the technical scale .


Prompt:
Some candidate values differ from the evidence by only one number. Select only the strings that appear verbatim, preserving candidate order. Return JSON using exactly this schema: {"reported_values":["candidate string"]}. Candidates: ["74.7%", "91.7%", "95%", "66.8%", "94%", "90.7%", "67.8%", "73.7%"]

Evidence:
Xylooligosaccharides (XOS) were produced from xylan-rich agricultural residues using endo-xylanase from Thermomyces lanuginosus PC7S1T immobilized on magnetic chitosan beads. The immobilized enzyme showed high immobilization yield (90.7%), efficiency (73.7%), and activity recovery (66.8%). Enhanced stability was observed, with 94% residual activity after 50 d at 4 °C and 54% after 10 reuse cycles, along with improved thermal (70–75 °C) and pH stability (pH 6.5). Immobilization altered the product profile, favoring xylotriose (X3) and xylotetraose (X4), whereas the free enzyme predominantly produced xylobiose (X2), highlighting its potential for prebiotic applications. Scanning electron microscopy confirmed effective substrate depolymerization. Toxicity assays using Artemia salina showed no significant differences in survival between XOS produced by free and immobilized enzymes ( p > 0.05), indicating low acute toxicity. Overall, these results reveal the potential of this immobilized system for sustainable and scalable production of prebiotic XOS from renewable biomass. Keywords: Xylooligosaccharides, Immobilized xylanase, Magnetic chitosan, Agricultural residues, Prebiotics


Prompt:
Identify every candidate statement reproduced exactly from the evidence. Return JSON using exactly this schema: {"supported":["A"]}. Preserve candidate order. JSON whitespace is ignored.

Evidence:
Multidrug resistance (MDR) remains the principal impediment to curative oncology, driven by complex interplays between cancer cells and the tumor microenvironment (TME). While nanomedicines have sought to overcome these delivery barriers, their clinical translation is often hampered by the heterogeneity of the enhanced permeability and retention (EPR) effect and by inefficient intratumoral delivery. In this review, we argue that overcoming MDR requires a transition beyond traditional passive drug delivery, advocating active, localized remodeling of the tumor ecosystem. Next-generation injectable hydrogels are increasingly recognized as localized viscoelastic niches that combine controlled intratumoral retention with the capacity to actively modulate biological responses within tumor TME. By converging principles of mechanobiology and immunometabolism, these hydrogels enable a multi-tiered strategy to dismantle multidimensional MDR. This approach begins with the biomechanical softening of the extracellular matrix to decouple mechanotransduction driven by Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ), followed by the metabolic disruption of hypoxia-driven bioenergetics. Beyond the extracellular landscape, nanogel-enabled trafficking allows payloads to circumvent intracellular sequestration and efflux transporters, while immunomodulatory niches mobilize antitumor immunity through in situ vaccination and myeloid reprogramming. Finally, we evaluate the integration of artificial intelligence-driven design and patient-derived organoids as a technical bridge to reconcile laboratory ingenuity with clinical utility, aiming to transform the TME into a vulnerable therapeutic target. The online version contains supplementary material available at 10.1186/s12943-026-02660-3.

Candidates:
A. Next-generation injectable hydrogels are not increasingly recognized as localized viscoelastic niches that combine controlled intratumoral retention with the capacity to actively modulate biological responses within tumor TME.
B. Next-generation injectable hydrogels are increasingly recognized as localized viscoelastic niches that combine controlled intratumoral retention with the capacity to actively modulate biological responses within tumor TME.
C. Multidrug resistance (MDR) remains the principal impediment to curative oncology, driven by complex interplays between cancer cells and the tumor microenvironment (TME).
D. In this review, we argue that overcoming MDR requires a transition beyond traditional passive drug delivery, advocating active, localized remodeling of the tumor ecosystem.
E. The evidence does not state that in this review, we argue that overcoming MDR requires a transition beyond traditional passive drug delivery, advocating active, localized remodeling of the tumor ecosystem.
F. While nanomedicines have sought to overcome these delivery barriers, their clinical translation is not often hampered by the heterogeneity of the enhanced permeability and retention (EPR) effect and by inefficient intratumoral delivery.
G. While nanomedicines have sought to overcome these delivery barriers, their clinical translation is often hampered by the heterogeneity of the enhanced permeability and retention (EPR) effect and by inefficient intratumoral delivery.
H. The evidence does not state that multidrug resistance (MDR) remains the principal impediment to curative oncology, driven by complex interplays between cancer cells and the tumor microenvironment (TME).