Tissue distribution of oral vitamin B12 is influenced by B12 status and B12 form: an experimental study in rats
Linda S. Kornerup 0 1
Sergey N. Fedosov 0 1
Christian B. Juul 0 1
Eva Greibe 0 1
Christian W. Heegaard 0 1
Ebba Nexo 0 1
0 Department of Molecular Biology and Genetics, Aarhus University , Aarhus , Denmark
1 Department of Clinical Biochemistry, Aarhus University Hospital , Palle Juul-Jensens Boulevard 99 8200 Aarhus N , Denmark
Purpose Hydroxocobalamin (HOCbl) is the dominating Cbl form in food, whereas cyanocobalamin (CNCbl) is common in vitamin pills and oral supplements. This study compares single-dose absorption and distribution of oral HO[57Co]Cbl and CN[57Co]Cbl in Cbl-deficient and normal rats. Methods Male Wistar rats (7 weeks) were fed a 14-day diet with (n = 15) or without (n = 15) Cbl. We compared the uptakes of HO[57Co]Cbl (free or bound to bovine transcobalamin) and free CN[57Co]Cbl administered by gastric gavage (n = 5 in each diet group). Rats were sacrificed after 24 h. Blood, liver, kidney, brain, heart, spleen, intestines, skeletal muscle, 24-h urine and faeces were collected, and the content of [57Co]Cbl was measured. Endogenous Cbl in tissues and plasma was analysed by routine methods. Results Mean endogenous plasma-Cbl was sevenfold lower in deficient vs. normal rats (190 vs. 1330 pmol/L, p < 0.0001). Cbl depletion increased endogenous Cbl ratios (tissue/plasma = kin/kout) in all organs except for the kidney, where the ratio decreased considerably. Twenty-four-hour accumulation of labelled Cbl showed that HOCbl > CNCbl (liver) and CNCbl > HOCbl (brain, muscle and plasma).
Hydroxocobalamin; Cyanocobalamin; Intestinal absorption; Cobalamin deficiency; Kinetic modelling
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Vitamin B12 (cobalamin, Cbl) is essential for a normal
neurological function and formation of blood cells. The
vitamin is supplied via dietary animal products and
increasingly through food fortification or vitamin pills [1]. The
Cbl forms present in foods are the coenzymes methyl- and
5′-deoxyadenosyl-Cbl (MeCbl, AdoCbl). A brief exposure
to light converts both coenzymes to hydroxo-Cbl (HOCbl),
making HOCbl the ubiquitous food form of Cbl [2].
CyanoCbl (CNCbl) is chemically stable, and it is the predominant
Cbl form used in industrial Cbl products (food fortification
and vitamin pills) [3]. The general concept is that HOCbl
and CNCbl are comparable concerning their absorption and
tissue distribution patterns, while free Cbl is absorbed more
efficiently than food bound Cbl (for a review see ref. [3]).
Our recent data in normal rats challenge both of these
statements [4]. We compared 24-h oral absorption of CN[57Co]
Cbl (free or bound to bovine transcobalamin (TC), the Cbl
binding protein in milk) and free HO[57Co]Cbl. We found
no difference between TC-bound and free CN[57Co]Cbl, as
well as no difference in the total absorption levels of the
two Cbl forms. However, the liver HO[57Co]Cbl
accumulation was twice as high as the CN[57Co]Cbl accumulation.
Notably, this result challenges the concept that CN[57Co]
Cbl and HO[57Co]Cbl behave alike and thereby are of equal
value for treatment/prevention of Cbl deficiency.
The current study was undertaken to investigate whether
the observed differences in distribution of HOCbl and
CNCbl also were mirrored in other tissues than liver and
kidney and whether this distribution was dependent on Cbl
status. In addition, we wanted to explore whether the food
form of Cbl (HOCbl) was absorbed alike when
administered free or bound to bovine TC.
Materials and methods
Thirty male Wistar rats (Taconic Bioscience Inc.,
Denmark) were used for the experiments; 7 weeks old,
weighing approx. 200 g upon arrival to the animal facilities.
The rats were housed in pairs in standard cages
(Makrolon 1291 H type III H, 800 cm2, Tecniplast, Italy) with
free access to food and tap water. The room temperature
was 19–20 °C and the humidity 60% with a 12/12 h light/
dark cycle. Bedding material (asp chips, Tapvei, Finland)
and soft paper wool (LBS biotech, United Kingdom) were
changed daily. Rats were kept for 2 weeks, during which
half (n = 15) were randomized to a Cbl-deficient diet
(Altromin C1024, Brogaarden, Denmark) and the other half
(n = 15) to the control diet (Altromin C1000, Brogaarden,
Denmark). The calorie contents of the two diets were equal,
but the Cbl-deficient diet contained less cellulose and corn
starch and more sucrose compared with the control diet.
The manufacturer assessed Cbl content by using the
tabulated values for Cbl in different food sources. Therefore, we
quantified Cbl in the diets by extracting 0.3 g of solids with
1.5 mL of water. After centrifugation, Cbl was measured in
the supernatant employing a Cobas 6000 (Roche
Diagnostics). During the analysis, all Cbl is converted to CNCbl;
thus, the Cbl content was calculated employing the
molecular weight for CNCbl, MW: 1355. The mean of two
independent measures is shown (Cbl-deficient diet: <0.5 (<0.5,
<0.5) µg/kg, control diet: 60 (69, 51) µg/kg).
All experiments were conducted in agreement with EU
Directive 2010/63/E (...truncated)