Serum Methylmalonic Acid Concentrations Research Articles

Aging-accumulated methylmalonic acid serum levels at breast cancer diagnosis are not associated with distant metastases.

Recent evidence suggests that age-accumulated methylmalonic acid (MMA) promotes breast cancer progression in mice. This study aims to investigate the association between baseline serum MMA concentrations in patients with breast cancer and the development of subsequent distant metastases. We included 32 patients with early Luminal B-like breast cancer (LumB, median age 62.4y) and 52 patients with early triple-negative breast cancer (TNBC, median age 50.5y) who developed distant metastases within 5years. They were matched to an equal number of early breast cancer patients (median age 62.2y for LumB and 50.5y for TNBC) who did not develop distant metastases with at least 5years of follow-up. Baseline serum MMA levels at breast cancer diagnosis showed a positive correlation with age (P < 0.001) and a negative correlation with renal function and vitamin B12 (all P < 0.02), but no statistical association was found with BMI or tumor stage (P > 0.6). Between matched pairs, no significant difference was observed in MMA levels, after adjusting for kidney function and age (P = 0.19). Additionally, in a mouse model, a significant decline in MMA levels was observed in the tumor-bearing group compared to the group without tumors before and after tumor establishment or at identical times for the control group (P = 0.03). Baseline serum MMA levels in patients with breast cancer are not correlated with secondary distant metastasis. Evidence in the mouse model suggests that the presence of a tumor perturbates MMA levels.

Efficacy and tolerance of oral versus parenteral cyanocobalamin supplement in hypocobalaminaemic dogs with chronic enteropathy: a controlled randomised open-label trial.

Determine comparative tolerance of daily oral and weekly parenteral cobalamin supplementation, in hypocobalaminaemic dogs with chronic enteropathy. Determine whether oral is as effective as parenteral supplementation at achieving eucobalaminaemia, in hypocobalaminaemic dogs with protein-losing enteropathy, severe hypocobalaminaemia or high canine inflammatory bowel disease activity index at inclusion. Thirty-seven client-owned dogs with hypocobalaminaemia and clinical signs of chronic enteropathy were prospectively enrolled in three UK referral centres. Dogs were randomly allocated to daily oral for 12 weeks or weekly parenteral cobalamin supplementation for 6 weeks and one additional dose 4 weeks later. Serum cobalamin, body condition score, canine inflammatory bowel disease activity index and bodyweight were assessed at inclusion, weeks 7 and 13. Serum methylmalonic acid concentration was evaluated at inclusion and at week 13. Owners completed treatment adherence, palatability, tolerance and satisfaction questionnaires at week 13. Nineteen dogs completed the study. All dogs orally supplemented achieved normal or increased cobalaminaemia at weeks 7 and 13. There was no statistical difference in cobalamin concentration at week 13 in dogs treated with oral or parenteral supplementation, regardless of presence of protein-losing enteropathy, severity of hypocobalaminaemia or canine inflammatory bowel disease activity index at inclusion. Serum methylmalonic acid concentration was not significantly different between oral and parenteral groups, neither were treatment adherence, satisfaction, and tolerance scores at week 13. Oral is as effective and as well-tolerated as parenteral cobalamin supplementation in hypocobalaminaemic dogs with chronic enteropathy and severe clinical or biochemical phenotypes, and should be considered as a suitable treatment option regardless of disease severity.

Serum methylmalonic acid concentrations at breast cancer diagnosis significantly correlate with clinical frailty.

Methylmalonic acid (MMA), a by-product of propionate metabolism, is known to increase with age. This study investigates the potential of serum MMA concentrations as a biomarker for age-related clinical frailty in older patients with breast cancer. One hundred nineteen patients ≥ 70years old with early-stage breast cancer were included (median age 76years). G8 screening, full geriatric assessment, clinical parameters (i.e., estimated glomerular filtration rate (eGFR) and body mass index (BMI)), and serum sample collection were collected at breast cancer diagnosis before any therapy was administered. MMA concentrations were measured via liquid chromatography with tandem mass spectrometry. MMA concentrations significantly increased with age and eGFR (all P < 0.001) in this older population. The group with an abnormal G8 (≤ 14, 51% of patients) had significantly higher MMA levels than the group with normal G8 (> 14, 49%): 260nmol/L vs. 188nmol/L, respectively (P = 0.0004), even after correcting for age and eGFR (P = 0.001). Furthermore, in the detailed assessment, MMA concentrations correlated most with mobility (Eastern Cooperative Oncology Group (ECOG) Performance Status and Activities of Daily Living (ADL) tools, all P ≤ 0.02), comorbidity (Charlson Comorbidity Index (CCI) tool, P = 0.005), and polypharmacy (P < 0.001), whereas no significant associations were noted for instrumental ADL (IADL), Mini-Mental State Examination (MMSE), Geriatric Depression Scale-15 (GDS15), Mini Nutritional Assessment-Short Form (MNA-SF), and pain (all P > 0.1). In addition, our results showed that higher MMA levels correlate with poor overall survival in breast cancer patients (P = 0.003). Elevated serum MMA concentrations at initial diagnosis are significantly associated, not only with age but also independently with clinical frailty, suggesting a possible influence of MMA on clinical frailty in older patients with early-stage breast cancer.

Serum cobalamin and methylmalonic acid concentrations in juvenile dogs with parvoviral enteritis or other acute enteropathies.

Low serum cobalamin concentrations have been associated with ileal malabsorption in dogs with chronic enteropathy. Increased serum methylmalonic acid (MMA) concentrations indicate cobalamin deficiency on a cellular level. Few studies have evaluated serum cobalamin concentrations or methylmalonic acid concentrations in juvenile dogs with parvoviral enteritis or nonparvoviral acute enteropathies. Evaluate serum cobalamin and methylmalonic acid concentrations in juvenile dogs (6 weeks to 10 months old) with parvoviral enteritis or nonparvoviral acute enteropathy. Thirty-one juvenile dogs with parvoviral enteritis, 29 dogs with nonparvoviral acute diarrhea (NPVAD), and 40 healthy juvenile control dogs. Single-center, prospective, observational, cross-sectional study. Serum cobalamin and, when sufficient serum was available, MMA concentrations were measured. Most serum cobalamin concentrations were within the adult reference interval. Serum cobalamin concentrations in healthy dogs (median, 848 ng/L; range, 293-1912 ng/L) were significantly higher than in dogs with parvoviral enteritis (P = .0002; median, 463 ng/L; range, <150-10 000 ng/L) or dogs with NPVAD (P = .02; median, 528 ng/L; range, 160-8998 ng/L). Serum MMA concentrations were not significantly different between groups (healthy dogs: median, 796 nmol/L; range, 427-1933 nmol/L; parvoviral enteritis: median, 858 nmol/L; range, 554-3424 nmol/L; NPVAD: median, 764 nmol/L; range, 392-1222 nmol/L; P = .1). Juvenile dogs with parvoviral enteritis or NPVAD had lower serum cobalamin concentrations than healthy juvenile dogs. However, based on serum MMA concentrations cellular cobalamin deficiency was not apparent.

Vitamin B12 in Cats: Nutrition, Metabolism, and Disease.

Cobalamin is a water-soluble molecule that has an important role in cellular metabolism, especially in DNA synthesis, methylation, and mitochondrial metabolism. Cobalamin is bound by intrinsic factor (IF) and absorbed in the ileal tract. The IF in cats is synthesized exclusively by pancreatic tissue. About 75% of the total plasma cobalamin in cats is associated with transcobalamin II, while in this species, transcobalamin I is not present. In cats, the half-life of cobalamin is 11-14 days. Diagnostic biomarkers for B12 status in cats include decreased levels of circulating total cobalamin and increased levels of methylmalonic acid. The reference interval for serum cobalamin concentrations in cats is 290-1500 ng/L, and for the serum methylmalonic acid concentration, it is 139-897 nmol/L. Therapy for hypocobalaminemia mainly depends on the underlying disease. In some cases, subcutaneous or intramuscular injection of 250 μg/cat is empirically administered. In recent years, it has been demonstrated that oral cobalamin supplementation can also be used successfully in dogs and cats as a less invasive alternative to parental administration. This review describes the current knowledge regarding B12 requirements and highlights improvements in diagnostic methods as well as the role of hypocobalaminemia in its associated diseases.

Abstract P2-11-18: Serum methylmalonic acid concentrations at breast cancer diagnosis are not associated with distant metastases

Abstract Introduction: Methylmalonic acid (MMA), a metabolite and by-product of propionate metabolism, promotes breast cancer (BC) progression in mice via the transforming growth factor-beta (TGFβ) signaling pathway (Gomes et al, Nature 2020). It is currently unknown if this effect also exists in patients with BC. Objectives: To investigate the association between baseline serum MMA concentrations in patients at BC diagnosis and development of distant metastases via a matched case-control study. Methods: We included 32 patients with early Luminal B-like BC (Lumb, median age 62.4y) and 52 patients with early triple-negative BC (TNBC, median age 50.5y) who developed distant metastases within 5 years. They were matched to an equal number of early BC patients with at least 5 years of follow-up (median age 62.2y for Lumb and 50.5y for TNBC) who did not develop distant metastases with at least 5 years of follow-up. Matching was performed based on age at diagnosis date (± 5y), tumor stage, and treatment received ((neo)adjuvant chemotherapy and radiotherapy, yes/no). Serum MMA concentrations were determined by liquid chromatography with tandem mass spectrometry (LC-MS-MS). Summary statistics, paired analyses, and multiple conditional logistic regression analyses were performed with and without adjusting for potential covariates (age, kidney function, and tumor stage). Results: Baseline serum MMA at BC diagnosis significantly correlated with age (rs=0.35, p=.005 in Lumb; rs=0.35, p=.0003 in TNBC), and negatively correlated with kidney function assessed by estimated glomerular filtration rate (eGFR, rs= -0.42, p=.0005 in Lumb; rs= -0.32, p=.0009 in TNBC). MMA concentrations at diagnosis were not associated with distant metastases in either subtype, after adjusting for kidney function, age, and tumor stage (all p&amp;gt;.05). Next, we categorized BC cases in the public TCGA (n=174 for Lumb; n=140 for TNBC), METABRIC (n=461 for Lumb; n=199 for TNBC), and GSE25066 (n=78 for Lumb; n=182 for TNBC) database according to their 5-year metastatic status, and analyzed the TGFβ signaling pathway activity of primary BC. Like MMA concentrations, a gene expression signature of TGFβ signaling was not associated with distant metastases in patients with BC. Conclusion: Baseline serum MMA concentrations and a gene signature for TGFβ signaling at BC diagnosis are not associated with distant metastases among patients with Lumb and TNBC subtypes. Citation Format: Qi Wu, Sigrid Hatse, Juan F. García, Patricia Altea-Manzano, Jaak Billen, Mélanie Planque, Anke Vandekeere, Yentl Lambrechts, François Richard, Annouschka Laenen, Kevin Punie, Patrick Neven, Ines Nevelsteen, Giuseppe Floris, Christine Desmedt, Ana Gomes, Sarah-Maria Fendt, Hans Wildiers. Serum methylmalonic acid concentrations at breast cancer diagnosis are not associated with distant metastases [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P2-11-18.

Abstract PD6-03: PD6-03 Serum methylmalonic acid concentrations at breast cancer diagnosis strongly correlate with frailty: a retrospective cross-sectional study

Abstract Introduction: Frailty commonly occurs in older persons, including those with breast cancer diagnoses. Methylmalonic acid (MMA), a metabolite and by-product of propionate metabolism, is known to increase significantly with aging. The relation between MMA concentrations and frailty is currently unknown. Objectives: A cross-sectional study was performed to study the association between baseline serum MMA concentrations and clinical frailty (estimated by G8 screening) in older patients with newly diagnosed breast cancer. Methods: 163 patients ≥70 years old with early-stage breast cancer were included (median age 76y). G8 screening and serum sample collection were performed at breast cancer diagnosis before any therapy was administered. MMA concentrations were measured via liquid chromatography with tandem mass spectrometry (LC-MS-MS). Results: MMA concentrations significantly increased with age (rs=0.3, p&amp;lt;.0001) and serum creatinine levels (rs=0.5, p&amp;lt;.0001) in this older population. The group with an abnormal G8 (≤14/17 = ‘frail’, 48% of patients) had significantly higher MMA levels than the group with normal G8 (&amp;gt;14/17 = ‘fit’, 52%): 250nM vs. 189 nM, respectively (p=.0002). Higher MMA concentrations were independently associated with abnormal G8 (Odds ratio, 1.003, 95%CI 1.0 to 1.006, p=.04) after adjusting for age and serum creatinine levels. Among the different components of G8, MMA concentrations correlated most with weight loss (rs= -0.18, p=.02), mobility (rs= -0.23, p=.002), and polypharmacy (rs= -0.22, p=.005). Conclusion: Elevated serum MMA concentrations at breast cancer diagnosis are significantly associated, not only with age but also independently with clinical frailty in older patients with early-stage breast cancer. MMA may be further evaluated as a biomarker of frailty in older persons with breast cancer. Citation Format: Qi Wu, Sigrid Hatse, Cindy Kenis, Yentl Lambrechts, Kevin Punie, Patrick Neven, Ann Smeets, Annouschka Laenen, Ana Gomes, Sarah-Maria Fendt, Hans Wildiers. PD6-03 Serum methylmalonic acid concentrations at breast cancer diagnosis strongly correlate with frailty: a retrospective cross-sectional study [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD6-03.

Correlations between circulating methylmalonic acid levels and all-cause and cause-specific mortality among patients with diabetes.

Evidence regarding serum methylmalonic acid (MMA) levels and mortality in individuals with diabetes is limited. This study aimed to evaluate the correlation between MMA and all-cause and cause-specific deaths in patients with diabetes. This is a population-based cohort study based on data from both the National Health and Nutrition Examination Survey (NHANES) and National Death Index from 1999 to 2014. We assessed the association of serum MMA concentrations with mortality using Cox proportional hazard models after adjusting for lifestyle, demographic factors, and comorbidities. Among the 3,097 participants, 843 mortalities occurred during a median follow-up of 4.42 years. There were 242 deaths due to cardiovascular disease (CVD) and 131 cancer-associated deaths. After multivariate adjustment, elevated serum MMA levels were markedly correlated with a high risk of all-cause, CVD-, and cancer-related deaths. Each one-unit increase in the natural log-transformed MMA level correlated with increased risk of all-cause mortality (2.652 times), CVD mortality risk (3.153 times), and cancer-related mortality risk (4.514). Hazard ratios (95% confidence intervals [CIs]) after comparing participants with MMA < 120 and ≥250 nmol/L were 2.177 (1.421-3.336) for all-cause mortality, 3.560 (1.809-7.004) for CVD mortality, and 4.244 (1.537-11.721) for cancer mortality. Higher serum MMA levels were significantly associated with higher all-cause, CVD, and cancer mortality. These findings suggest that maintaining lower MMA status may lower mortality risk in individuals with diabetes.

233 (PB-057) Poster - Serum methylmalonic acid concentrations at breast cancer diagnosis are not associated with distant metastases

Background: Methylmalonic acid (MMA), a metabolite and by-product of propionate metabolism, promotes breast cancer (BC) progression in mice via the transforming growth factor beta (TGFβ) signaling pathway (Gomes et al, Nature 2020). It is currently unknown if this effect also exists in patients with BC. This study is to investigate the association between baseline serum MMA concentrations in patients at BC diagnosis and the development of distant metastases via a matched case-control study.

Effect of oral or injectable supplementation with cobalamin in dogs with hypocobalaminemia caused by chronic enteropathy or exocrine pancreatic insufficiency.

BackgroundRecent studies have shown similar efficacy of oral supplementation of cobalamin compared to injectable supplementation in dogs, but few prospective, randomized studies have been published.ObjectivesTo evaluate efficacy of oral or injectable supplementation with cobalamin in normalizing serum cobalamin and methylmalonic acid (MMA) concentrations in dogs with hypocobalaminemia caused by either chronic enteropathy (CE) or exocrine pancreatic insufficiency (EPI).AnimalsForty‐six client owned dogs with hypocobalaminemia.MethodsProspective randomized clinical trial. Dogs were divided into 2 groups (CE or EPI), and randomized to receive oral or injectable supplementation of cobalamin. Each dog had 3 visits and serum cobalamin and MMA concentrations were measured at each visit.ResultsIn dogs with CE, serum cobalamin concentrations increased with oral (P = .02; median 149 [range 149‐231] to 733 [166‐1467] ng/L, median difference 552 [95% CI: 181‐899] ng/L) or injectable (P < .01; 168 [149‐233] to 563 [234‐965] ng/L, 367 [187‐623] ng/L) supplementation. In dogs with EPI, serum cobalamin concentrations increased with oral (P = .01; 162 [149‐214] to 919 [643‐3863] ng/L, 705 [503‐3356] ng/L) or injectable (P = .01; 177 [149‐217] to 390 [243‐907] ng/L, 192 [89‐361] ng/L) supplementation. Serum MMA concentrations decreased with oral or injectable supplementation in dogs with CE, but only with oral supplementation in dogs with EPI.Conclusions and Clinical ImportanceOral supplementation is an alternative for cobalamin supplementation in dogs with hypocobalaminemia caused by CE or EPI.

High-throughput analysis of total homocysteine and methylmalonic acid with the efficiency to separate succinic acid in serum and urine via liquid chromatography tandem mass spectrometry

Vitamin B12 (VB12) deficiency may lead to hyperhomocysteinemia and methylmalonic acidemia development which are risk factors of cardiovascular disease and nervous system impairment, respectively. However, few analytical methods are available to simultaneously quantify total homocysteine (tHcy) and methylmalonic acid (MMA) due to complex analytical requirements, such as sensitivity at nanomolar concentration, separation performance for succinic acid (SA), an endogenous isomer of MMA, and retention properties for polar compounds. Therefore, we developed and validated a simple and accurate liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of tHcy and MMA with the efficient separation of SA in human serum and urine. The clinical performance of the assay was validated according to CLSI C62-A guidelines. The recovery for serum tHcy was 95.2–105.8%, urine tHcy was 98.1–111.5%, serum MMA was 94.6–99.4%, and urine MMA was 101.6–105.6%. In addition, the LC-MS/MS method was found to be reliable based on the value of inter-assay imprecision and total imprecision coefficient variation (CV), matrix effect, and carryover. Standards and samples were stable in −20 °C for at least 2 months. The limits of quantifications (LOQs) were 0.074 nmol/mL for tHcy and 0.040 nmol/mL for MMA, which are suitable for detecting tHcy and MMA concentrations in human serum and urine. The concentration of tHcy and MMA in samples collected from 148 subjects were measured using this method. The results suggested that the concentrations of serum tHcy and MMA considerably differed between VB12 sufficient and deficient groups. Serum tHcy and serum MMA concentrations were inversely correlated with VB12 status. Our method represents a rapid technique for estimating tHcy and MMA concentrations in serum and urine samples without the need for derivatization and may be used to assess VB12 status in clinical applications.

The role of vitamin B12 in viral infections: a comprehensive review of its relationship with the muscle-gut-brain axis and implications for SARS-CoV-2 infection.

This comprehensive review establishes the role of vitamin B12 as adjunct therapy for viral infections in the treatment and persistent symptoms of COVID-19, focusing on symptoms related to the muscle–gut–brain axis. Vitamin B12 can help balance immune responses to better fight viral infections. Furthermore, data from randomized clinical trials and meta-analysis indicate that vitamin B12 in the forms of methylcobalamin and cyanocobalamin may increase serum vitamin B12 levels, and resulted in decreased serum methylmalonic acid and homocysteine concentrations, and decreased pain intensity, memory loss, and impaired concentration. Among studies, there is much variation in vitamin B12 doses, chemical forms, supplementation time, and administration routes. Larger randomized clinical trials of vitamin B12 supplementation and analysis of markers such as total vitamin B12, holotranscobalamin, total homocysteine and methylmalonic acid, total folic acid, and, if possible, polymorphisms and methylation of genes need to be conducted with people with and without COVID-19 or who have had COVID-19 to facilitate the proper vitamin B12 form to be administered in individual treatment.

1-13C-propionate breath testing as a surrogate endpoint to assess efficacy of liver-directed therapies in methylmalonic acidemia (MMA)

PurposeTo develop a safe and noninvasive in vivo assay of hepatic propionate oxidative capacity. MethodsA modified 1-13C-propionate breath test was administered to 57 methylmalonic acidemia (MMA) subjects, including 19 transplant recipients, and 16 healthy volunteers. Isotopomer enrichment (13CO2/12CO2) was measured in exhaled breath after an enteral bolus of sodium-1-13C-propionate, and normalized for CO2 production. 1-13C-propionate oxidation was then correlated with clinical, laboratory, and imaging parameters collected via a dedicated natural history protocol. ResultsLower propionate oxidation was observed in patients with the severe mut0 and cblB subtypes of MMA, but was near normal in those with the cblA and mut− forms of the disorder. Liver transplant recipients demonstrated complete restoration of 1-13C-propionate oxidation to control levels. 1-13C-propionate oxidation correlated with cognitive test result, growth indices, bone mineral density, renal function, and serum biomarkers. Test repeatability was robust in controls and in MMA subjects (mean coefficient of variation 6.9% and 12.8%, respectively), despite widely variable serum methylmalonic acid concentrations in the patients. ConclusionPropionate oxidative capacity, as measured with 1-13C-propionate breath testing, predicts disease severity and clinical outcomes, and could be used to assess the therapeutic effects of liver-targeted genomic therapies for MMA and related disorders of propionate metabolism. TRIAL REGISTRATIONThis clinical study is registered in www.clinicaltrials.gov with the ID: NCT00078078. Study URL: http://clinicaltrials.gov/ct2/show/NCT00078078 [Display omitted]

Anemia, iron deficiency, and cobalamin deficiency in cats with chronic gastrointestinal disease.

BackgroundIron deficiency and cobalamin deficiency, as sequelae to chronic gastrointestinal (GI) disease, could result in anemia and increased morbidity in cats with chronic enteropathies.ObjectiveTo evaluate iron deficiency in cats with chronic GI disease and its relationship with hypocobalaminemia, anemia, and disease severity.AnimalsTwenty client‐owned cats with primary GI disease.MethodsProspective, cross‐sectional study. Cats were enrolled at the time of evaluation for chronic GI disease, after exclusion of comorbidities. CBC with reticulocyte indices, iron metabolism (serum iron and ferritin concentrations, total iron binding capacity [TIBC]), serum methylmalonic acid (MMA), cobalamin, and folate concentrations, pancreatic lipase and trypsin‐like immunoreactivity, and disease severity were evaluated.ResultsAnemia (hematocrit <30%), iron deficiency, and cobalamin deficiency were diagnosed in 4/20, 7/20, and 8/20 cats, respectively. Hematocrit (r s = −.45; P < .05) and body condition score (r s = −.60; P < .01) negatively correlated with MMA. Median TIBC was lower in cats with increased vs normal MMA (218 μg/mL; range, 120‐466 μg/mL vs 288 μg/mL; range, 195‐369 μg/mL; P = .02). Hematocrit (r s = .51; P = .02), reticulocyte MCV (r s = .52; P = .02), reticulocyte hemoglobin content (r s = .71; P < .001), and percent transferrin saturation (r s = .79; P < .0001) positively correlated with serum iron concentration.Conclusions and Clinical ImportanceFunctional iron deficiency was common in cats with chronic GI disease. Associations between hypocobalaminemia, iron parameters, and hematologic parameters warrant further investigation on the impact of iron deficiency on chronic GI disease morbidity in cats.

Efficacy of intramuscular hydroxocobalamin supplementation in cats with cobalamin deficiency and gastrointestinal disease.

BackgroundIn humans, absorption and tissue retention rates of intramuscularly administered hydroxocobalamin (OH‐Cbl) are superior compared to cyanocobalamin (CN‐Cbl). Supplementation with OH‐Cbl has not been described in cats.ObjectivesTo evaluate effects of parenteral OH‐Cbl supplementation on clinical signs, serum Cbl and methylmalonic acid (MMA) concentrations in hypocobalaminemic cats with gastrointestinal disease.AnimalsTwenty‐three client‐owned cats.MethodsProspective study. Serum Cbl and MMA concentrations were determined at enrollment (t0), immediately before the 4th OH‐Cbl IM injection (300 μg, given q2 weeks) (t1), and 4 weeks after the 4th injection (t2). Severity of clinical signs (activity, appetite, vomiting, diarrhea, body weight) was graded at each time point and expressed as clinical disease activity score.ResultsMedian clinical disease activity score decreased significantly from t0 (6; range, 2‐10) to t1 (1; range, 0‐6) and t2 (1; range, 0‐9). Median serum Cbl concentration increased significantly from 111 pmol/L (range, 111‐218; reference range, 225‐1451 pmol/L) at t0 to 1612 pmol/L (range, 526‐14 756) (P < .001) at t1, and decreased again significantly to 712 pmol/L (range, 205‐4265) (P < .01) at t2. Median baseline serum MMA concentration at t0 (802 nmol/L; range, 238‐151 000; reference range, 120‐420 nmol/L) decreased significantly (P < .001) to 199 nmol/L (range, 29‐478) at t1, and was 205 nmol/L (range, 88‐734) at t2. Serum MMA concentrations normalized in 22/23 cats at t1, and were not significantly higher at t2 compared to t1.Conclusions and Clinical ImportanceThe herein described OH‐Cbl injection scheme appears efficacious for normalization of cellular Cbl deficiency in cats with gastrointestinal disease.

Assessment of folate and cobalamin concentrations in relation to their dependent intracellular metabolites in serum of pigs between 6 and 26 weeks of age

Folate (vitamin B9) and cobalamin (vitamin B12) play an important role in amino acid metabolism, nucleic acid synthesis, and methyl group transfer. Two intracellular enzymes, methionine synthase and methylmalonyl-CoA mutase, are folate and/or cobalamin-dependent, respectively. At the cellular level, a lack of folate and cobalamin leads to accumulation of serum homocysteine (HCY) and a lack of cobalamin leads to increased methylmalonic acid (MMA) concentrations. Altered serum HCY and MMA concentrations can influence amino acid metabolism and nucleic acid synthesis in pigs. Therefore, we aimed to evaluate serum folate, cobalamin, HCY, and MMA concentrations in postweaning pigs between 6 and 26 weeks of age. Serum samples from 12 pigs collected at week 6, 7, 8, 9, 10, 14, 18, 22, and 26 as part of an unrelated study were analyzed. Serum folate (p < .0001), cobalamin (p = .0001), HCY (p < .0001), and MMA (p < .0001) concentrations differed significantly during the postweaning period between 6 and 26 weeks of age; with significantly higher serum HCY (at weeks 6 and 7 compared to weeks 9, 14, 18, 22, and 26) and MMA concentrations (at weeks 6, 7, and 8 compared to weeks 14, 18, 22, and 26) and an overall decrease of serum MMA concentrations from week 6 to week 14 in the pigs studied. This study suggests age-dependent changes in intracellular folate- and cobalamin-dependent metabolites (i.e., HCY and MMA) in pigs between 6 and 26 weeks of age, possibly reflecting decreased availability of intracellular folate and/or cobalamin for amino acid metabolism, nucleic acid synthesis, and methyl group transfer.

Maternal choline supplementation alters vitamin B-12 status in human and murine pregnancy

Despite participation in overlapping metabolic pathways, the relationship between choline and vitamin B-12 has not been well characterized especially during pregnancy. We sought to determine the effects of maternal choline supplementation on vitamin B-12 status biomarkers in human and mouse pregnancy, hypothesizing that increased choline intake would improve vitamin B-12 status. Associations between common genetic variants in choline-metabolizing genes and vitamin B-12 status biomarkers were also explored in humans. Healthy third-trimester pregnant women (n=26) consumed either 480 or 930 mg choline/day as part of a 12-week controlled feeding study. Wild-type NSA and Dlx3 heterozygous (Dlx3+/−) mice, which display placental insufficiency, consumed a 1×, 2× or 4× choline diet and were sacrificed at gestational days 15.5 and 18.5. Serum vitamin B-12, methylmalonic acid (MMA) and homocysteine were measured in all samples; holotranscobalamin (in humans) and hepatic vitamin B-12 (in mice) were also measured. The 2× choline supplementation for 12 weeks in pregnant women yielded higher serum concentrations of holotranscobalamin, the bioactive form of vitamin B-12 (~24%, P=.01). Women with genetic variants in choline dehydrogenase (CHDH) and betaine-homocysteine S-methyltransferase (BHMT) had higher serum MMA concentrations (~31%, P=.03) and lower serum holotranscobalamin concentrations (~34%, P=.03), respectively. The 4× choline dose decreased serum homocysteine concentrations in both NSA and Dlx3+/− mice (~36% and~43% respectively, P≤.015). In conclusion, differences in choline supply due to supplementation or genetic variation modulate vitamin B-12 status during pregnancy, supporting a functional relationship between these nutrients.

Age-specific reference ranges are needed to interpret serum methylmalonic acid concentrations in the US population

Serum vitamin B-12 is measured to evaluate vitamin B-12 status. Serum methylmalonic acid (MMA) is a specific functional indicator of vitamin B-12 status; however, concentrations increase with impaired renal function. The aim of this study was to describe the distribution of serum vitamin B-12 and MMA in US adults, and estimate age-specific reference intervals for serum MMA in a healthy subpopulation with replete vitamin B-12 status and normal renal function. We examined cross-sectional data for serum vitamin B-12 and MMA in adults participating in the NHANES from 2011 to 2014. Vitamin B-12 was measured by electrochemiluminescence assay and MMA by isotope-dilution liquid chromatography-tandem mass spectrometry. In both bivariate and multivariate analyses, age, race/Hispanic origin, and vitamin B-12 supplement use were generally significantly associated with serum vitamin B-12 and MMA concentrations. Serum MMA concentrations increased with age, particularly in persons aged ≥70 y. Non-Hispanic white persons had lower vitamin B-12 and higher MMA concentrations than non-Hispanic black persons. Shorter fasting times and impaired renal function were significantly associated with higher serum MMA concentrations, but not with serum vitamin B-12 concentrations after controlling for covariates. The central 95% reference intervals for serum vitamin B-12 and MMA concentrations were widest for persons aged ≥70 y compared with younger age groups. Compared with the overall population, the central 95% reference intervals for serum MMA concentrations were considerably narrower for a vitamin B-12-replete subpopulation with normal renal function, but still age-dependent. Serum vitamin B-12 showed little, whereas serum MMA showed notable, increases with impaired renal function. The higher serum MMA concentrations throughout the entire distribution in older persons (especially persons aged ≥70 y) who are vitamin B-12-replete and have normal renal function indicate the need for age-specific MMA reference intervals to better interpret vitamin B-12 status in epidemiologic research.

Age-specific Serum Methylmalonic Acid Reference Ranges for More Accurate Interpretation of Vitamin B-12 Status in U.S. Adults (P24-030-19)

Abstract Objectives To evaluate vitamin B-12 status of U.S. adults. High serum methylmalonic acid (MMA) is a specific functional indicator of low vitamin B-12 status, but it is not sensitive because concentrations also increase with impaired renal function. Our goal was to describe the distribution of serum vitamin B-12 and MMA in U.S. adults, and estimate age-specific reference intervals for serum MMA in a subpopulation with replete vitamin B-12 status and normal renal function. Methods We examined cross-sectional data for serum vitamin B-12 and MMA in U.S. adults participating in the National Health and Nutrition Examination Survey from 2011–2014. Vitamin B-12 was assessed by electrochemiluminescence assay and MMA by isotope-dilution LC-MS/MS. Results In both bivariate and multivariate analysis age, race/Hispanic origin, and vitamin B-12 supplementation were generally significantly associated with serum vitamin B-12 and MMA concentrations before and after controlling for additional covariates. We observed higher serum vitamin B-12 and MMA concentrations in persons ≥60 y compared with younger age groups. Non-Hispanic white persons had lower vitamin B-12 and higher MMA concentrations compared with non-Hispanic black persons. Shorter fasting times and impaired renal function were significantly associated with higher serum MMA concentrations, but not with serum vitamin B-12 concentrations after controlling for covariates. The central 95% reference intervals for serum vitamin B-12 and MMA concentrations were wider for persons ≥60 y compared with younger age groups. Compared with the overall population, the central 95% reference interval for serum MMA concentrations was considerably narrower for a vitamin B-12 replete subpopulation with normal renal function, but still age-dependent. Serum vitamin B-12 showed little, while serum MMA showed notable increases with impaired renal function. Conclusions The higher serum MMA concentrations throughout the entire distribution in older persons who are vitamin B-12 replete and have normal renal function indicate the need for age-specific MMA reference intervals to better interpret the vitamin B-12 status in clinical and epidemiologic research. Funding Sources This work was supported by direct appropriations from U.S. Congress.

Reference intervals for serum total vitamin B12 and holotranscobalamin concentrations and their change points with methylmalonic acid concentration to assess vitamin B12 status during early and mid-pregnancy.

Background Maternal vitamin B12 (B-12) adequacy is important for maternal health and optimal fetal growth. However, pregnancy-specific cut-offs for B-12 biomarkers are lacking. Methods Reference intervals for serum total B-12, holotranscobalamin (holoTC) and methylmalonic acid (MMA) concentrations were calculated following CLSI EP28-A3c guidelines in 723 pregnant women of European (50%) and South Asian (50%) ethnicity, residing in British Columbia, Canada, at median (range) 11.4 (8.3-13.9) and 16.1 (14.9-20.9) weeks of gestation. Change point analyses described relationships between log serum MMA concentration with serum total B-12 and holoTC concentrations, assuming linear-linear relationships. Results The central 95% reference interval limits indicated that serum total B-12 <89.9 and <84.0 pmol/L, holoTC <29.5 and <26.0 pmol/L and MMA >371 and >374 nmol/L, in the first and second trimesters, respectively, may indicate B-12 deficiency in pregnant women. The lower limits of total B-12 and holoTC and the upper limits of MMA significantly differed by ethnicity in both trimesters. According to the change point analysis, total B-12 <186 and <180 pmol/L and holoTC <62.2 and <67.5 pmol/L in the first and second trimesters, respectively, suggested an increased probability of impaired intracellular B-12 status, with no difference between ethnicities. Conclusions We present novel reference limits and change points for B-12 biomarkers, which may be employed to identify possible B-12 deficiency in women during early and mid-pregnancy. Future research is needed to validate these cut-offs and determine the predictors and functional outcomes associated with impaired B-12 status in ethnically diverse populations.