Structural basis of respiratory complex adaptation to cold temperatures.

In response to cold, mammals activate brown fat for respiratory-dependent thermogenesis reliant on the electron transport chain. Yet, the structural basis of respiratory complex adaptation upon cold exposure remains elusive. Herein, we combined thermoregulatory physiology and cryoelectron microscopy (cryo-EM) to study endogenous respiratory supercomplexes from mice exposed to different temperatures. A cold-induced conformation of CI:III2 (termed type 2) supercomplex was identified with a ∼25° rotation of CIII2 around its inter-dimer axis, shortening inter-complex Q exchange space, and exhibiting catalytic states that favor electron transfer. Large-scale supercomplex simulations in mitochondrial membranes reveal how lipid-protein arrangements stabilize type 2 complexes to enhance catalytic activity. Together, our cryo-EM studies, multiscale simulations, and biochemical analyses unveil the thermoregulatory mechanisms and dynamics of increased respiratory capacity in brown fat at the structural and energetic level.

Magnolol from Magnolia officinalis inhibits Neopestalotiopsis ellipsospora by damaging the cell membrane.

Tea gray blight disease is a significant threat to the tea industry. In this study, a biological activity approach was utilized to investigate the efficacy of green fungicides from Magnolia officinalis stem bark against Neopestalotiopsis ellipsospora. The active compounds were isolated and purified, and their structures were elucidated. In vitro and in vivo activity screenings revealed that the n-hexane extract, which contained magnolol and honokiol, exhibited strong activity against N. ellipsospora, showing complete inhibition at 100 mg/L. The EC50 values of magnolol and honokiol were 5.11 and 6.09 mg/L, respectively. Mechanistically, magnolol was found to disrupt N. ellipsospora invasion by damaging the cell membrane, increasing permeability, and causing leakage of intracellular substances. Transcriptome analysis revealed that magnolol treatment downregulates membrane-related genes and leads to the enrichment of lipid metabolism pathway genes. This study revealed that magnolol inhibits N. ellipsospora growth by affecting lipid metabolism and compromising cell membrane integrity.

Determining the Effect of Non-Thermal Plasma on the Transmembrane Kinetics of Melittin through Molecular Explorations.

Non-thermal plasma (NTP) synergistic anticancer strategies are a current hotspot of interest at the intersection of plasma biomedicine. Melittin (MEL) has been shown to inhibit cancer in many malignant tumors; however, its clinical application is controversial. Therefore, the transmembrane process and mechanism of MEL activity in different cell systems were studied and the combination of MEL and NTP was proposed in this paper. The results showed that the electrostatic attraction between MEL and the lipid bilayer contributes to the stable orientation of MEL on the membrane surface. In addition, sialic acid overexpression affects the degree to which MEL binds the membrane system and the stability of the membrane structure. The use of NTP to reduce the dosage of MEL and its related nonspecific cytolysis activity has certain clinical application value. The results of this study provide theoretical support for improving the clinical applicability of MEL and contribute to the further development of plasma biomedicine.

Heat shock factor ZmHsf17 positively regulates phosphatidic acid phosphohydrolase ZmPAH1 and enhances maize thermotolerance.

Heat stress (HS) adversely impacts plant growth, development and grain yield. Heat shock factors (Hsf), especially HsfA2 subclass, play a pivotal role in the transcriptional regulation of genes in response to HS. In this study, the coding sequence of maize ZmHsf17 was cloned. ZmHsf17 contains conserved domains: DNA binding, oligomerization and transcriptional activation. The protein was nuclear localized and had transcription activation activity. Yeast two hybrid and split luciferase complementary assays confirmed the interaction of ZmHsf17 with members of the maize HsfA2 subclass. Overexpression of ZmHsf17 in maize significantly increased chlorophyll content and net photosynthesis rate of maize leaves, and enhanced the stability of cellular membranes. Through integrative analysis of ChIP-seq and RNA-seq datasets, ZmPAH1, encoding phosphatidic acid phosphohydrolase of lipid metabolic pathways, was identified as a target gene of ZmHsf17. The promoter fragment of ZmPAH1 was bound by ZmHsf17 in protein-DNA interaction experiments in vivo and in vitro. Lipidomic data also indicates that the overexpression of ZmHsf17 increased levels of some critical membrane lipid components of maize leaves under HS. This research provides new insights into the role of the ZmHsf17-ZmPAH1 module in regulating thermotolerance in maize.

Biochemical Pathways Delivering Distinct Glycosphingolipid Patterns in MDA-MB-231 and MCF-7 Breast Cancer Cells.

The complex structure of glycosphingolipids (GSLs) supports their important role in cell function as modulators of growth factor receptors and glutamine transporters in plasma membranes. The aberrant composition of clustered GSLs within signaling platforms, so-called lipid rafts, inevitably leads to tumorigenesis due to disturbed growth factor signal transduction and excessive uptake of glutamine and other molecules needed for increased energy and structural molecule cell supply. GSLs are also involved in plasma membrane processes such as cell adhesion, and their transition converts cells from epithelial to mesenchymal with features required for cell migration and metastasis. Glutamine activates the mechanistic target of rapamycin complex 1 (mTORC1), resulting in nucleotide synthesis and proliferation. In addition, glutamine contributes to the cancer stem cell GD2 ganglioside-positive phenotype in the triple-negative breast cancer cell line MDA-MB-231. Thieno[2,3-b]pyridine derivative possesses higher cytotoxicity against MDA-MB-231 than against MCF-7 cells and induces a shift to aerobic metabolism and a decrease in S(6)nLc4Cer GSL-positive cancer stem cells in the MDA-MB-231 cell line. In this review, we discuss findings in MDA-MB-231, MCF-7, and other breast cancer cell lines concerning their differences in growth factor receptors and recent knowledge of the main biochemical pathways delivering distinct glycosphingolipid patterns during tumorigenesis and therapy.

Hydrogen sulfide attenuates chilling injury in loquat fruit by alleviating oxidative stress and maintaining cell membrane integrity.

The effects of hydrogen sulfide (H2S) on chilling injury (CI), reactive oxygen species (ROS) metabolism, sugar metabolism, pentose phosphate pathway (PPP), and membrane lipid metabolism in loquat fruit throughout the refrigerated period were investigated in this study. The findings indicated that H2S application restrained the increase in internal browning (IB), malondialdehyde (MDA) content, and electrolyte leakage, while sustaining higher total phenolic and total flavonoid levels, and lower soluble quinone content in loquat fruit. Besides, H2S promoted antioxidant accumulation and increased antioxidant enzyme activities by the regulation of ROS metabolism, along with increasing fructose and glucose levels and reducing power by activating sugar metabolism and PPP. Furthermore, H2S treatment retarded the degradation of phospholipids and fatty acids in loquat fruit by modulating membrane lipid metabolism relevant enzyme activities. These findings indicated that H2S application mitigated CI in loquat fruit by alleviating oxidative stress and maintaining cell membrane structural integrity.

Exogenous nitric oxide treatment delays the senescence of postharvest mung bean sprouts by regulating ascorbic acid metabolism.

BACKGROUND: This study evaluated the effects of nitric oxide (NO) treatment on ascorbic acid (AsA) metabolism and mung bean sprout quality. It examined changes in the AsA content, enzyme activity associated with AsA metabolism, antioxidant capacity, cell membrane composition, and cellular structure to clarify the effects of NO on mung bean sprouts. RESULTS: Nitric oxide treatment preserved mung bean sprout quality by enhancing significantly the activity of enzymes involved in the l-galactose pathway (including guanosine diphosphate (GDP)glutathione (-d-mannose pyrophosphorylase, GDP-mannose-3',5'-epimerase, GDP-l-galactose phosphorylase, l-galactose-1-phosphate phosphatase, l-galactose dehydrogenase, and l-galactose-1,4-lactone dehydrogenase) and the AsA-glutathione (GSH)(Beijing Solarbio Science and Technology Co.,Ltd., Beijing, China) cycle (including ascorbate peroxidase, ascorbic acid oxidase, glutathione reductase, dehydroascorbate reductase, and monodehydroascorbate reductase) during the germination and storage stage. Increased enzyme activity led to an increase in AsA content and enhanced antioxidant capacity, and reduced the membrane lipid damage in mung bean sprouts. This was demonstrated by higher levels of DPPH radical scavenging capacity, unsaturated fatty acids and phospholipids, along with lower levels of hydrogen peroxide, superoxide anions, and malondiadehyde, in NO-treated mung bean sprouts. Scanning electron microscopy also revealed that NO treatment maintained the integrity of the cellular structure of the mung bean sprouts. CONCLUSION: Nitric oxide accelerates AsA metabolism effectively by regulating the biosynthesis and regeneration of AsA in mung bean sprouts. These changes increased AsA levels, alleviated membrane lipid damage, delayed senescence, and maintained the quality of mung bean sprouts during storage. © 2024 Society of Chemical Industry.

Phospholipid biosynthesis regulation for improving pigment production by Monascus in response to ammonium chloride stress.

In the actual industrial production process, the efficient biosynthesis and secretion of Monascus pigments (MPs) tend to take place under abiotic stresses, which often result in an imbalance of cell homeostasis. The present study aimed to thoroughly describe the changes in lipid profiles in Monascus purpureus by absolute quantitative lipidomics and tandem mass tag-based quantitative proteomics. The results showed that ammonium chloride stress (15 g/L) increased MP production while inhibiting ergosterol biosynthesis, leading to an imbalance in membrane lipid homeostasis in Monascus. In response to the imbalance of lipid homeostasis, the regulation mechanism of phospholipids in Monascus was implemented, including the inhibition of lysophospholipids production, maintenance of the ratio of PC/PE, and improvement of the biosynthesis of phosphatidylglycerol, phosphatidylserine, and cardiolipin with high saturated and long carbon chain fatty acids through the CDP-DG pathway rather than the Kennedy pathway. The inhibition of lysophospholipid biosynthesis was attributed to the upregulated expression of protein and its gene related to lysophospholipase NTE1, while maintenance of the PC/PE ratio was achieved by the upregulated expression of protein and its gene related to CTP: phosphoethanolamine cytidylyltransferase and phosphatidylethanolamine N-methyltransferase in the Kennedy pathway. These findings provide insights into the regulation mechanism of MP biosynthesis from new perspectives.IMPORTANCEMonascus is important in food microbiology as it produces natural colorants known as Monascus pigments (MPs). The industrial production of MPs has been achieved by liquid fermentation, in which the nitrogen source (especially ammonium chloride) is a key nutritional parameter. Previous studies have investigated the regulatory mechanisms of substance and energy metabolism, as well as the cross-protective mechanisms in Monascus in response to ammonium chloride stress. Our research in this work demonstrated that ammonium chloride stress also caused an imbalance of membrane lipid homeostasis in Monascus due to the inhibition of ergosterol biosynthesis. We found that the regulation mechanism of phospholipids in Monascus was implemented, including inhibition of lysophospholipids production, maintenance of the ratio of PC/PE, and improvement of biosynthesis of phosphatidylglycerol, phosphatidylserine, and cardiolipin with high saturated and long carbon chain fatty acids through the CDP-DG pathway. These findings further refine the regulatory mechanisms of MP production and secretion.

Maintaining quality of postharvest green pepper fruit using melatonin by regulating membrane lipid metabolism and enhancing antioxidant capacity.

Green pepper quality often deteriorates during storage because of membrane lipid damage and oxidative stress. This study investigated the effects of exogenous melatonin (MT) on green pepper storage quality, membrane lipids, and antioxidant metabolism. The results showed that MT increased the activities of superoxide dismutase, catalase, ascorbate peroxidase, peroxidase, monodehydroascorbate reductase, and dehydroascorbate reductase in green peppers compared to the control group. It upregulated expression of multiple enzymes; reduced accumulation of reactive oxygen species such as dehydroascorbic acid, H2O2, and O2.-; and maintained high ascorbic acid, glutathione, coenzyme II, and nicotinamide adenine dinucleotide while reducing oxidized glutathione levels. In addition, MT decreased lipoxygenase and phospholipase D activities, downregulated ReLOX and RePLD expression, and delayed the degradation of phosphatidylcholine, phosphatidylethanolamine, and oleic, linoleic, and linolenic acids in green peppers. These results suggest that MT helps to improve the chilling injury and quality of green peppers and extends shelf life.

Neurolipidomics in schizophrenia: A not so well-oiled machine.

Most patients with schizophrenia (SCZ) do not exhibit violent behaviors and are more likely to be victims rather than perpetrators of violent acts. However, a subgroup of forensic detainees with SCZ exhibit tendencies to engage in criminal violations. Although numerous models have been proposed, ranging from substance use, serotonin transporter gene, and cognitive dysfunction, the molecular underpinnings of violence in SCZ patients remains elusive. Lithium and clozapine have established anti-aggression properties and recent studies have linked low cholesterol levels and ultraviolet (UV) radiation with human aggression, while vitamin D3 reduces violent behaviors. A recent study found that vitamin D3, omega-3 fatty acids, magnesium, and zinc lower aggression in forensic population. In this review article, we take a closer look at aryl hydrocarbon receptor (AhR) and the dysfunctional lipidome in neuronal membranes, with emphasis on cholesterol and vitamin D3 depletion, as sources of aggressive behavior. We also discuss modalities to increase the fluidity of neuronal double layer via membrane lipid replacement (MLR) and natural or synthetic compounds. This article is part of the Special Issue on "Personality Disorders".

Slightly acidic electrolyzed water treatment enhances the quality attributes and the storability of postharvest litchis through regulating the metabolism of reactive oxygen species.

Effects of slightly acidic electrolyzed water (SAEW) on the storability, quality attributes, and reactive oxygen species (ROS) metabolism of litchis were investigated. Results showed that SAEW-treated litchis presented better quality attributes and storability than control litchis. On storage day 5, the commercially acceptable fruit rate of control litchis was 42%, while SAEW-treated litchis displayed 59% higher rate of commercially acceptable fruit, 21% lower pericarp browning index, and 13% lower weight loss percentage than control litchis. Additionally, compared to control litchis, SAEW-treated litchis demonstrated higher activities of SOD, CAT and APX, higher levels of GSH, AsA, DPPH radical scavenging ability, and reducing power, but lower O2 -· generation rate, lower levels of H2O2 and MDA. These findings indicated that SAEW treatment could elevate antioxidant capacity and ROS scavenging ability, reduce ROS production and accumulation, and lower membrane lipid peroxidation, thereby retaining the quality attributes and storability of litchis.

Defect in Migration of HSPCs in Nox-2 Deficient Mice Explained by Impaired Activation of Nlrp3 Inflammasome and Impaired Formation of Membrane Lipid Rafts.

NADPH oxidase 2 (Nox2), a superoxide-generating enzyme, is a source of reactive oxygen species (ROS) that regulate the intracellular redox state, self-renewal, and fate of hematopoietic stem/progenitor cells (HSPCs). Nox2 complex expressed on HSPCs associated with several activated cell membrane receptors increases the intracellular level of ROS. In addition, ROS are also released from mitochondria and, all together, are potent activators of intracellular pattern recognition receptor Nlrp3 inflammasome, which regulates the trafficking, proliferation, and metabolism of HSPCs. In the current study, we noticed that Nox2-deficient mice, despite the increased number of HSPCs in the bone marrow (BM), show hematopoietic defects illustrated by delayed recovery of peripheral blood (PB) hematopoietic parameters after sublethal irradiation and mobilize fewer HSPCs after administration of G-CSF and AMD3100. Moreover, Nox2-deficient HSPCs engraft poorly after transplantation into normal syngeneic recipients. To explain these defects at the molecular level, we hypothesized that Nox2-KO decreased ROS level does not efficiently activate Nlrp3 inflammasome, which plays a crucial role in regulating the trafficking of HSPCs. Herein, we report Nox2-deficient HSPCs display i) defective migration to major chemoattractant, ii) impaired intracellular activation of Nlrp3 inflammasome, and iii) a defect in membrane lipid raft (MLRs) formation that is required for a proper chemotactic response to pro-migratory factors. We conclude that Nox2-derived ROS enhances in Nlrp3 inflammasome-dependent manner HSPCs trafficking by facilitating MLRs assemble on the outer cell membranes, and defect in Nox2 expression results in impaired activation of Nlrp3 inflammasome, which affects HSPCs migration.

A lipid synthase maintains metabolic flux for jasmonate synthesis to regulate root growth and phosphate homeostasis.

Plants require phosphate (Pi) for proper growth and development but often face scarcity of this vital nutrient in the soil. Pi-starvation triggers membrane lipid remodeling to utilize the membrane phospholipid-bound Pi in plants. In this process, phospholipids are replaced by non-Pi-containing galactolipids (MGDG, DGDG) and sulfolipids. The galactolipids ratio (MGDG:DGDG) is suggested to influence jasmonic acid (JA) biosynthesis. However, how the MGDG:DGDG ratio, JA levels, and root growth are coordinated under Pi deficiency in rice (Oryza sativa) remains unknown. Here, we characterized DGDG synthase 1 (OsDGD1) for its role in regulating root development by maintaining metabolic flux for JA biosynthesis. We showed that OsDGD1 is responsive under low Pi and is under the direct control of Phosphate Starvation Response 2 (OsPHR2), the master regulator of low Pi adaptations. Further, OsDGD1 knockout (KO) lines showed marked phenotypic differences compared to the wild type (WT), including a significant reduction in root length and biomass, leading to reduced Pi uptake. Further, lipidome analyses revealed reduced DGDG levels in the KO line, leading to reduced membrane remodeling, thus affecting P utilization efficiency. We also observed an increase in the MGDG: DGDG ratio in KO lines, which enhanced the endogenous JA levels and signaling. This imbalance of JA in KO plants led to changes in auxin levels, causing drastic root growth inhibition. These findings indicate the critical role of OsDGD1 in maintaining optimum levels of JA during Pi deficiency for conducive root growth. Besides acting as signaling molecules and structural components, our study widens the role of lipids as metabolic flux controllers for phytohormone biosynthesis.

Investigating the role of membrane lipid composition differences on spray drying survival in Lactobacillus bulgaricus using non-targeted Lipidomics.

The cell membrane, consisting of a phospholipid bilayer, is an important defense system of lactic acid bacteria (LAB) against adverse conditions. However, this membrane gets damaged during the process of spray drying of LAB into powder. In this study, two strains of Lactobacillus bulgaricus L9-7 and L4-2-12 with significantly different survival rates of about 22.49% and 0.43% after spray drying were explored at the cell membrane level. A total of 65 significantly different lipid species were screened from the cell membranes of two strains, with cardiolipin (CL) 15:1_22:6_24:0_28:0 being the crucial lipid species affecting membrane resistance. Finally, the KEGG enrichment analysis revealed that glycerophospholipid metabolism was the most predominant pathway, and eleven lipid species were annotated, including CL. Overall, this paper provides valuable insights into enhancing the heat tolerance of LAB.

γ-aminobutyric acid treatment inhibits browning and promotes storage quality by regulating reactive oxygen species and membrane lipid metabolism in fresh-cut stem lettuce.

The effects of γ-aminobutyric (GABA) on enzymatic browning, storage quality, membrane and reactive oxygen species (ROS) metabolism in fresh-cut stem lettuce were investigated. The results illustrated that GABA treatment delayed browning degree, polyphenol oxidase (PPO) activity and the expression of LsPPO. Meanwhile, higher chlorophyll and ascorbic acid contents were exhibited in GABA-treated stem lettuce, as well as the slower microbial propagation. Further investigation revealed that exogenous GABA application declined malondialdehyde content, electrolyte leakage and the enzyme activities of membrane metabolism, and the expression levels of related genes were also downregulated. In addition, GABA treatment scavenged ROS and strengthened the enzyme activities of ROS metabolism, as well as the expression levels of corresponding genes. Taken together, these findings implied that the repressed enzymatic browning and microbial propagation in GABA-treated stem lettuce were due to the inhibition of ROS accumulation, enhancement of membrane stability and increased resistance to oxidation.

Positive residues of the SARS-CoV-2 fusion domain are key contributors to the initiation of membrane fusion.

SARS-CoV-2 is one of the most infectious viruses ever recorded. Despite a plethora of research over the last several years, the viral life cycle is still not well understood, particularly membrane fusion. This process is initiated by the fusion domain (FD), a highly conserved stretch of amino acids consisting of a fusion peptide (FP) and fusion loop (FL), which in synergy perturbs the target cells' lipid membrane to lower the energetic cost necessary for fusion. In this study, through a mutagenesis-based approach, we have investigated the basic residues within the FD (K825, K835, R847, K854) utilizing an in vitro fusion assay and 19F NMR, validated by traditional 13C 15N techniques. Alanine and charge-conserving mutants revealed every basic residue plays a highly specific role within the mechanism of initiating fusion. Intriguingly, K825A led to increased fusogenecity which was found to be correlated to the number of amino acids within helix one, further implicating the role of this specific helix within the FD's fusion mechanism. This work has found basic residues to be important within the FDs fusion mechanism and highlights K825A, a specific mutation made within the FD of the SARS-CoV-2 spike protein, as requiring further investigation due to its potential to contribute to a more virulent strain of SARS-CoV-2.

A novel inducible animal model for studying chronic plasmalogen deficiency associated with Alzheimer's disease.

Plasmalogens are vinyl-ether glycerophospholipids critical for the structure and function of neuronal membranes. Deficient plasmalogen levels are associated with neurodegenerative diseases, particularly Alzheimer's disease (AD), which has led to the hypothesis that plasmalogen deficiency might drive disease onset and progression. However, the lack of a suitable animal model with late-onset plasmalogen deficiency has prevented testing of this hypothesis. The goal of this project was therefore to develop and characterize a mouse model capable of undergoing a plasmalogen deficiency only in adulthood, mirroring the chronic decline thought to occur in AD. We report here the creation of a novel animal model containing a tamoxifen-inducible knockout of the Gnpat gene encoding the first step in the plasmalogen biosynthetic pathway. Tamoxifen treatment in adult animals resulted in a significant reduction of plasmalogens in both the circulation and tissues as early as four weeks. By four months, changes in behavior and nerve function were observed, with strong correlations between residual brain plasmalogen levels, hyperactivity, and latency. The model will be useful for further elucidating the role of plasmalogens in AD and evaluating plasmalogen therapies.

Yet more evidence that non-aqueous myelin lipids can be directly imaged with ultrashort echo time (UTE) MRI on a clinical 3T scanner: a lyophilized red blood cell membrane lipid study.

Direct imaging of semi-solid lipids, such as myelin, is of great interest as a noninvasive biomarker of neurodegenerative diseases. Yet, the short T2 relaxation times of semi-solid lipid protons hamper direct detection through conventional magnetic resonance imaging (MRI) pulse sequences. In this study, we examined whether a three-dimensional ultrashort echo time (3D UTE) sequence can directly acquire signals from membrane lipids. Membrane lipids from red blood cells (RBC) were collected from commercially available blood as a general model of the myelin lipid bilayer and subjected to D2O exchange and freeze-drying for complete water removal. Sufficiently high MR signals were detected with the 3D UTE sequence, which showed an ultrashort T2* of ∼77-271 µs and a short T1 of ∼189 ms for semi-solid RBC membrane lipids. These measurements can guide designing UTE-based sequences for direct in vivo imaging of membrane lipids.

OsMGD1-Mediated Membrane Lipid Remodeling Improves Salt Tolerance in Rice.

Salt stress severely reduces photosynthetic efficiency, resulting in adverse effects on crop growth and yield production. Two key thylakoid membrane lipid components, monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), were perturbed under salt stress. MGDG synthase 1 (MGD1) is one of the key enzymes for the synthesis of these galactolipids. To investigate the function of OsMGD1 in response to salt stress, the OsMGD1 overexpression (OE) and RNA interference (Ri) rice lines, and a wild type (WT), were used. Compared with WT, the OE lines showed higher chlorophyll content and biomass under salt stress. Besides this, the OE plants showed improved photosynthetic performance, including light absorption, energy transfer, and carbon fixation. Notably, the net photosynthetic rate and effective quantum yield of photosystem II in the OE lines increased by 27.5% and 25.8%, respectively, compared to the WT. Further analysis showed that the overexpression of OsMGD1 alleviated the negative effects of salt stress on photosynthetic membranes and oxidative defense by adjusting membrane lipid composition and fatty acid levels. In summary, OsMGD1-mediated membrane lipid remodeling enhanced salt tolerance in rice by maintaining membrane stability and optimizing photosynthetic efficiency.

Phototherapy Alters the Plasma Metabolite Profile in Infants Born Preterm with Hyperbilirubinemia.

OBJECTIVE: To investigate the effects of gestational age (GA) and phototherapy on the plasma metabolite profile of preterm infants with neonatal hyperbilirubinemia (NHB). STUDY DESIGN: From a cohort of prospectively enrolled infants born preterm (n = 92), plasma samples of very preterm (VPT; GA, 28 + 0 to 31 + 6 weeks, n = 27) and moderate/late preterm (M/LPT; GA, 32 + 0 to 35 + 6 weeks, n = 33) infants requiring phototherapy for NHB were collected prior to the initiation of phototherapy and 24 hours after starting phototherapy. An additional sample was collected 48 hours after starting phototherapy in a randomly selected subset (n = 30; VPT n = 15; M/LPT n = 15). Metabolite profiles were determined using ultraperformance liquid chromatography tandem mass spectroscopy. Two-way ANCOVA was used to identify metabolites that differed between GA groups and timepoints after adjusting for total serum bilirubin levels (false discovery rate q-value < 0.05). Top impacted pathways were identified using pathway over-representation analysis. RESULTS: Phototherapy was initiated at lower total serum bilirubin (mean ± SD mg/dL) levels in VPT compared with M/LPT infants (7.3 ± 1.4 vs 9.9 ± 1.9, P < .01). We identified 664 metabolites that were significant for a phototherapy effect, 191 metabolites significant for GA, and 46 metabolites significant for GA × phototherapy interaction (false discovery rate q-value < 0.05). Longer duration phototherapy had a larger mean effect size (24 hours postphototherapy: d = 0.36; 48 hours postphototherapy: d = 0.43). Top pathways affected by phototherapy included membrane lipid metabolism, one-carbon metabolism, creatine biosynthesis, and oligodendrocyte differentiation. CONCLUSION: Phototherapy alters the plasma metabolite profile more than GA in preterm infants with NHB, affecting pathways related to lipid and one-carbon metabolism, energy biosynthesis, and oligodendrocyte differentiation.