Supplementary Digital Content (SDC)

S1: Screening evaluation in all patients

Etiological Evaluation

◦ Hepatotrophic viruses- IgM Hepatitis A, IgM Hepatitis E, HbsAg, IgM Anti

Hbc.

Other viruses- IgM Epstein Barr virus, IgM Cytomegalovirus, IgM

Parvo virus (where indicated).

◦ Wilson disease-Serum Ceruloplasmin, 24 hour urinary copper, DCT, Evidence

of Hemolysis, KF Ring (If possible).

◦ Autoimmune- DCT, ANA/ASMA/LKM-1/SLA, IgG.

◦ Hemophagocytosis- Serum triglyceride, cholesterol, ferritin and bone marrow

biopsy (If Fever/Splenomegaly/Bicytopenia)

Subject Evaluation

Detailed historical evaluation including age, gender, clinical

symptomatology, detailed past, development, birth and family history was

elicited from the subjects/parents.

Laboratory evaluation

All the subjects underwent detailed lab evaluation including complete liver

function test (serum bilirubin, aspartate aminotransferase, alanine

aminotransferase, alkaline phosphatase, gamma glutamyl transpeptidase,

albumin), international normalized ratio, serum creatinine, blood urea, serum

electrolytes, serum uric acid, metabolic Screen (random plasma glucose,

arterial ammonia, arterial blood gas analysis, urinary ketones, urine non

glucose reducing substances, serum creatine phosphokinase).

Echocardiography and Ultrasonography of abdomen was performed by trained

and experienced operators. Samples for metabolic testing included plasma

sample for Tandem Mass Spectrometry (TMS) for Carnitine/Acylcarnitine

Profile (and amino-acid analysis) and urine sample for Gas Chromatography

and Mass Spectrometry (GCMS) were collected within 4 hours of admission.

Age wise cut offs were used for analyzing metabolites’ results [1-4]

S2: Specific metabolic abnormalities detected in fatty acid oxidation disorders

Fatty acid oxidation disorder Plasma acylcarnitine Urine organic acidsCarnitine uptake defect Decreased C0 (free

carnitine), Decreased long-chainacylcarnitines

Normal or Elevated dicarboxylic acids

Carnitine palmitoyltransferase-I Elevated C0, Decreased long-chain acylcarnitines

Usually normal

Carnitine palmitoyltransferase-II Elevated C16, C18:1, C14, low C2

Normal

Carnitine acylcarnitine translocase Elevated C16, C18:1, C14, low C2

Normal or Elevated dicarboxylic acids

Very-long-chain acyl-CoAdehydrogenase

Elevated C14, C14:1/C12:1

Elevated Dicarboxylic acids

Long-chain 3-hydroxyacyl-CoAdehydrogenase/mitochondrialtrifunctional protein

Elevated C16 OH, C18:1 OH, C14 OH

Elevated Dicarboxylic acids, Elevated hydroxydicarboxylic acids (3-hydroxyadipic acid, 3-hydroxysebacic acid)

Medium-chain ketoacyl-CoA thiolase

Unknown Elevated Lactic, 3-hydroxybutyric, saturated and unsaturated C6–C16dicarboxylic acids

Multiple acyl-CoA dehydrogenase/glutaric aciduria II

Elevated C4, C5, C5 DC, C6, C8, C10, C12, C14

Elevated Glutaric acid, isobutyrylglycine, ethylmalonic acid,dicarboxylic acids, acylglycines (phenylpropionylglycine),2-hydroxyglutaric acid

Succinyl-CoA: oxoacid transferase Normal NormalMedium-chain acyl-CoAdehydrogenase

Elevated C8, C10, C10:1 Elevated Dicarboxylic acids, acylglycines (suberylglycine,hexanoylglycine), 5-hydroxyhexanoic acid, octanedioic acid, decanedioic acid

Medium/short-chain acyl-CoAdehydrogenase

Elevated 3-OH-C4 Elevated Dicarboxylic acids, 2-hydroxyglutaric acid

Short-chain acyl-CoA dehydrogenase

Elevated C4 (butyrylcarnitine and butyrylglycine)

Elevated Ethylmalonate, methylsuccinate

Ethylmalonic encephalopathy Elevated C4, C5 Elevated Lactic acid, ethylmalonate, methylsuccinate

2,4-Dienoyl reductase Hypocarnitinemia, 2-trans, 4-cisdecadienoylcarnitinein both urine and blood

Normal

3-Hydroxy-3-methylglutaryl-CoAsynthase 2

Normal Normal

3-Hydroxy-3-methylglutaryl-CoAlyase

Elevated C5OH Elevated 3-Methylglutaric acid, 3-methylglutaconic acid, 3-hydroxy-3-methylglutaric acid, 3-methylcrotonylglycine

β-Ketothiolase Normal or Decreased C0 Tiglylglycine, acetoacetic acid, 2-methylacetoacetic acid

S3: Consort diagram depicting patient selection

Abbreviation: ALF- Acute liver failure

Group 1Children (0-18 years age)

with ALF(n= 38)

Group 2Adults (> 18 Years age) with ALF

(n=26)

Step 1History , Baseline and Etiological Evaluation

All ALF Cases

(n= 55)Pediatric Group(n= 33)

Adult Group(n = 22)

Step 2 Metabolic

Testing

Excluded:On Valproate Therapy (n=2)Delayed sampling beyond 4 hours (n = 3)

Excluded:Delayed sampling beyond 4 hours (n = 4)

S4- Comparison of proportions for predicting outcome (Death or Liver transplantation/LT vs Survival) in the overall group (n = 55)

Parameter(Normal Value)

Death/LT(n= 27)

Survived (n= 28)

OR, 95 % CI , p-Value

High Methionine 24 17 5.18, 1.25 to 21.41, 0.016

High Phenyalanine 8 1 11.37, 1.31 to 98.59, 0.012

High Tyrosine 15 6 4.58, 1.41 to 14.91, 0.009

High Arginine 4 2 2.26, 0.38 to 13.51, 0.422

Hyperaminoacidemia

(elevation of > 1 aminoacid)

24 19 3.789, 0.89 to 15.97, 0.059

Low Free Carnitine (C0) 12 11 1.236, 0.42 to 3.61, 0.698

Abnormal Carnitine profile

(any abnormality)

14 17 0.69, 0.24 to 2.03, 0.508

Abbreviations- OR- Odd’s Ratio, CI- Confidence interval.

S5: Liver biopsy figure showing Diffuse Hepatocyte steatosis with minimal fibrosis and inflammation in Case 1

S6: Liver biopsy figure showing Steatohepatitis with Pericellular fibrosis in Case 2

a b c

a b

References

1. Schmidt-Sommerfeld E, Werner E, Penn D: Carnitine plasma concentrations in 353

metabolically healthy children. Eur J Pediatr 1988;147:356-360.

2. http://geneticslab.emory.edu/support//AR_reference_ranges.pdf. Accessed January 2,

2015.

3. http://geneticslab.emory.edu/support//AA_reference_ranges.pdf. Accessed January 2,

2015.

4. http://geneticslab.emory.edu/support//OA_reference_ranges.pdf. Accessed January 2,

2015.