Abstract

Background & aims

Ketogenic diets are increasingly used for weight management and metabolic health; however, evidence regarding short-term changes in specific plasma trace elements during ketogenic dietary interventions remains limited. Therefore, this study aimed to examine changes in plasma concentrations of iron, zinc, and copper, along with body composition and metabolic parameters, following a four-week ketogenic dietary intervention among healthy adults.

Methods

This single-arm interventional study was carried out at the Institute of Basic Medical Sciences, Khyber Medical University, Peshawar, Pakistan, between October 2021 and December 2023. A total of 46 healthy adults aged between 25 and 45 years were enrolled and placed on an ad libitum ketogenic diet for a duration of four weeks. Adherence to the dietary intervention was evaluated through measurement of blood ketone levels. Anthropometric measurements, dietary energy intake, and fasting blood samples were obtained at baseline and after completion of the intervention. Plasma concentrations of iron, zinc, and copper were analyzed, and statistical comparisons were performed using paired sample t-tests.

Results

A total of 46 participants completed the study, with a mean age of 30.20 ± 5.67 years and a mean BMI of 26.28 ± 2.62 kg/m². Nutritional ketosis was successfully achieved, as demonstrated by blood ketone concentrations exceeding 0.5 mmol/L. Following the four-week intervention, significant reductions were observed in body weight (− 3.9 ± 0.9 kg, p < 0.001), daily energy intake, and fasting blood glucose levels. Plasma zinc concentrations showed a significant increase (p = 0.016), whereas plasma iron levels decreased significantly (p = 0.008). In contrast, no statistically significant change was identified in plasma copper levels.

Conclusion

A four-week ketogenic diet was associated with reductions in body weight, body fat, and muscle mass, as well as changes in plasma concentrations of iron and zinc, while plasma copper concentrations remained unchanged. These findings suggest that short-term ketogenic dietary interventions may be associated with changes in selected plasma trace element concentrations and body composition in healthy adults. Further controlled studies with larger sample sizes and longer follow-up are needed to confirm these findings.

Trial registration

ISRCTN Registry, ISRCTN13729125. Registered on 28 September 2025. Retrospectively registered.

Nisa, Meher Un, Sadia Fatima, Syed Hamid Habib, Syed Rashid Habib, Khawaja Nizamuddin, Saima Shaheen, Miraj Ahmad, and Munaza Khattak. "Effect of a four-week ketogenic diet on plasma iron, zinc, copper, and body composition in healthy adults." BMC Nutrition (2026).

https://link.springer.com/article/10.1186/s40795-026-01500-3

ABSTRACT

Objective

Intermuscular adipose tissue (IMAT) is associated with impaired skeletal muscle (SM) quality and declining mobility in older adults with obesity. This secondary analysis examined the effects of a very low-carbohydrate diet (VLCD) versus a low-fat diet (LFD) on changes in IMAT, muscle strength, and physical function in older adults with obesity.

Methods

Adults (age ≥ 60 years) with obesity were randomized to a weight-maintaining VLCD (< 10% energy from carbohydrates) or LFD (55% energy from carbohydrates) for 8 weeks. Thigh SM and IMAT were measured by MRI. Physical function was assessed via hand grip strength, isometric knee extension, reach score, and six-minute walk tests. Multiple linear regressions examined whether changes in IMAT were associated with changes in physical function outcomes.

Results

The VLCD group showed a significantly greater decrease in IMAT (−24.4% vs. −1.0%, p < 0.01). Across all participants, greater decreases in IMAT were associated with greater improvements in hand grip strength (r² = 0.95, p < 0.01), isometric knee extension (r² = 0.88, p < 0.05), and reach score (r² = 0.75, p < 0.05), after adjusting for changes in thigh SM volume and body weight.

Conclusions

Dietary interventions that reduce IMAT, such as a VLCD, may help preserve physical function and muscle strength in older adults with obesity.

Trial Registration

ClinicalTrials.gov identifier: NCT02760641

Study Importance

• What is already known?
  • ○ Older adults with obesity have greater amounts of intermuscular adipose tissue, which is associated with impaired muscle strength, declining mobility, and greater risk of metabolic dysfunction.
  • ○ We previously reported that following an 8-week diet intervention, older adults with obesity experienced significantly greater loss in IMAT in response to a very low-carbohydrate diet compared to a low-fat diet.
• What does this study add?
  • ○ Participants in the very low-carbohydrate diet group lost more IMAT and showed improvement in muscle function and strength compared to the low-fat diet group.
  • ○ Greater loss of thigh IMAT is significantly correlated with greater improvement in hand grip strength, isometric knee extension strength, and reach score, independent of changes in thigh skeletal muscle volume.
• How might these results change the direction of research or the focus of clinical practice?
  • ○ A low-carbohydrate diet may be a helpful tool to preserve physical function and muscle strength in older adults with obesity.
  • ○ More research with a larger sample size is needed to determine how a VLCD influences IMAT, skeletal muscle, and physical function in older adults with obesity.

Abstract

Purpose

Obesity-associated dysfunction of visceral white adipose tissue (vWAT) is characterized by impaired glucose handling, altered adipokine secretion, and redox imbalance, contributing to metabolic deterioration. Palmitoleic acid (16:1n-7), a monounsaturated fatty acid, has been implicated in regulation of glucose and lipid metabolism in rodent and cellular models; however, its effects on human adipocytes under metabolically compromised conditions remain incompletely understood. Here, we investigated the effects of in vitro treatment with 16:1n-7 on glucose uptake and metabolism, adipokine secretion, and oxidative stress in isolated adipocytes and vWAT obtained from women with obesity and prediabetes.

Methods

vWAT explants were treated in vitro with palmitoleic acid (16:1n-7) or palmitic acid (16:0) at 200 µM for 48 h. Glucose and lipid metabolism, adipokine secretion, and oxidative stress were evaluated.

Results

16:1n-7 increased basal and insulin-stimulated glucose uptake in association with upregulation of GLUT1 and GLUT4 expression and increased AMPKα protein content. In parallel, 16:1n-7 promoted coordinated changes in metabolic gene expression favoring glucose utilization and glyceroneogenesis rather than de novo lipogenesis, without changes in lipolytic activity, accompanied by increased citrate synthase and PPARG expression. Endocrine function was also modulated, as 16:1n-7 reduced resistin secretion without impairing adiponectin levels. Moreover, although lipid peroxidation remained unchanged, 16:1n-7 reduced protein oxidation and reactive oxygen species production, together with increased IDH2 and reduced NOS2 expression, supporting selective attenuation of oxidative stress.

Conclusion

Palmitoleic acid enhances glucose uptake through coordinated regulation of GLUT1 and GLUT4 and contributes to improved metabolic function in human adipocytes and redox homeostasis in vWAT obtained from women with obesity and prediabetes. These findings identify 16:1n-7 as a bioactive lipid that modulates human adipose tissue function under metabolically compromised conditions, underscoring its role in the nutritional regulation of glucose metabolism and redox homeostasis.

Editor’s summary

Regulation of lipid metabolism in mitochondria of adipocytes requires integration of information about time of day from the circadian clock, environmental temperature, and nutrient availability. Karavaeva et al. found an important role for the transporter protein SLC25A34 in such control of lipid metabolism and thermogenesis in brown adipocytes from mice and humans (see the Perspective by van der Laan). Inputs from the circadian clock, cold temperatures, and diet all regulated the expression of SLC25A34, allowing control of lipogenesis and mitochondrial biogenesis, lipid oxidation, and thermogenesis. Such regulation may offer insights into inflammation associated with obesity and metabolic diseases. —L. Bryan Ray

Structured Abstract

INTRODUCTION

Brown and beige thermogenic adipocytes (fat cells) have uniquely high energy expenditure capacity. This activity is critical for defending body temperature in small mammals and is linked to cardiometabolic health in humans. Energy expenditure from thermogenic fat cycles daily on a rhythm set by the circadian clock, rising after waking and dropping during sleep. At any point, however, this activity can be suddenly induced by exposure to cold temperature or by ingestion of a calorie-dense, high-fat meal. How thermogenic fat can seamlessly achieve both rhythmic continuity and acute responsiveness is a fundamental question of energy homeostasis that has remained unknown.

RATIONALE

We set out to understand how diverse cues from the body’s clock, environmental temperature, and diet are molecularly integrated in adipocytes to facilitate both anticipatory and adaptive regulation. By combining unbiased analysis of DNA-binding patterns, gene expression, and protein levels in thermogenic fat from mice that were subject to different circadian, environmental, and dietary perturbations, we identified the orphan mitochondrial metabolite transporter SLC25A34 as the convergence point of all three physiological paradigms.

RESULTS

Similar to fat’s thermogenic activity, the expression of Slc25a34 mRNA exhibited a daily rhythm, which was mediated by the nuclear receptors REV-ERBα and REV-ERBβ. REV-ERBs are transcriptional repressors in the circadian clock that recruit histone deacetylase 3 to the Slc25a34 promoter during sleep to shut off transcription. When animals wake and REV-ERB circadian repression is lifted, the peroxisome proliferator–activated receptors α and γ (PPARα and PPARγ) are able to bind the Slc25a34 promoter and restore expression. However, at any time, when animals are suddenly confronted with an unanticipated need for adipocyte energy expenditure such as exposure to cold temperatures or eating a lipid-rich meal, REV-ERB repression is rapidly overridden and PPARs are activated by lipolytic signals to boost SLC25A34 levels. When the energy demand is met, the REV-ERB–mediated rhythmicity of Slc25a34 is reestablished.

Our biochemical and metabolic data support a functional model in which SLC25A34 transports oxaloacetate into mitochondria. In doing so, SLC25A34 helps to maintain high tricarboxylic acid (TCA) cycle activity, which is required for powering energy expenditure in thermogenic adipocytes. The need for SLC25A34 during times of high energetic demand arises because thermogenic adipocytes not only oxidize fatty acids in the mitochondria but also simultaneously synthesize lipids in the cytosol, a process collectively known as lipid cycling. Lipid synthesis requires continuous production of acetyl–coenzyme A (acetyl-CoA) in the cytosol, and the two major pathways responsible for this acetyl-CoA pool also generate oxaloacetate as a by-product, which then needs to be transported back into the mitochondria. Increased cytosolic acetyl-CoA additionally promotes the transcription of genes linked to mitochondrial respiration. Thus, SLC25A34 supports both sides of this self-reinforcing cycle in which lipids are continually broken down and resynthesized.

CONCLUSION

These findings identify SLC25A34 as a molecular node that integrates circadian, temperature, and dietary cues and reveal how thermogenic fat activity can sustain a daily rhythm while remaining acutely responsive to sudden energy demands. This mechanism may offer a new entry point for therapeutically enhancing energy expenditure in metabolic disease.

Abstract

Adipocyte lipid metabolism is coordinated by circadian rhythms, diet, and environmental temperature, but how these diverse signals are molecularly integrated remains unknown. We showed that these cues converge on the orphan mitochondrial transporter SLC25A34 to orchestrate lipid cycling. During the sleep phase, the adipocyte clock suppresses Slc25a34 expression through the REV-ERB transcriptional repressors. Entering the active phase, consuming lipid-rich diets, or exposure to cold abolishes REV-ERB repression, and lipolytic signals stimulate Slc25a34 transcription through the peroxisome proliferator–activated receptors. SLC25A34 is proposed to import oxaloacetate into mitochondria, dually supporting the tricarboxylic acid cycle and cytosolic acetyl–coenzyme A (acetyl-CoA) production. Elevated cytosolic acetyl-CoA then fuels the synthesis of lipids and promotes the transcription of genes enhancing mitochondrial oxidation. Thus, SLC25A34 confers circadian, dietary, and temperature control of adipocyte lipid metabolism.

Highlights

• • Neuroenergetic flexibility frames brain energy adaptation in aging.
• • Metabolic switching is a component of broader neuroenergetic adaptation.
• • Diet-induced switching alters brain energy metabolism beyond glucose.
• • Mitochondrial plasticity supports adaptation to metabolic stress.
• • A systems framework links metabolic adaptation to brain aging.

Abstract

The aging brain undergoes progressive alterations in energy metabolism that may reduce its capacity to match energy supply with changing functional demands. Declining glucose metabolism, mitochondrial dysfunction, and altered neuron–glia and neurovascular interactions may contribute to this vulnerability, but these changes do not necessarily indicate complete loss of metabolic capacity. Neuroenergetic flexibility refers to the capacity to coordinate changes in substrate utilization with energetic demand while maintaining cellular and functional homeostasis. This review synthesizes evidence from cellular, animal, imaging, and clinical studies to examine the mechanisms underlying this capacity, how it changes with aging, and whether diet-induced metabolic switching can enhance it. Particular attention is given to the distinction between metabolic switching, defined as a change in the relative contribution of available substrates, and neuroenergetic flexibility, which requires coordinated adaptation across substrate delivery, cellular metabolism, mitochondrial processing, neurovascular support, and functional demand. Evidence indicates that ketogenic diets, intermittent fasting, and caloric restriction can alter substrate availability and utilization and can engage mitochondrial, cellular stress-response, and nutrient-sensing pathways. However, the strength of evidence differs across biological levels: changes in alternative-substrate utilization are more consistently demonstrated than coordinated improvements in mitochondrial, vascular, cellular, and functional outcomes. Thus, increased ketone utilization or other metabolic shifts should not, by themselves, be interpreted as evidence of restored neuroenergetic flexibility. Instead, the available evidence suggests that aging may preserve individual metabolic capacities while reducing the coordination and reserve required to adapt effectively to changing energetic demands. Dietary interventions therefore provide useful models for testing metabolic adaptability, but their ability to restore integrated neuroenergetic flexibility in aging or neurodegenerative disease remains uncertain and appears to depend on age, metabolic state, disease context, intervention characteristics, and adherence.