Homocoenzyme B12 and Bishomocoenzyme B12: Covalent Structural Mimics for Homolyzed,Enzyme‐Bound Coenzyme B12

Efficient electrochemical syntheses of “homocoenzyme B₁₂” ( 2 , Co_β‐(5′‐deoxy‐5′‐adenosyl‐methyl)‐cob(III )alamin) and “bishomocoenzyme B₁₂” ( 3 , Co_β‐[2‐(5′‐deoxy‐5′‐adenosyl)‐ethyl]‐cob(III )alamin) are reported here. These syntheses have provided crystalline samples of 2 and 3 in 94 and 77 % yield, respectively. In addition, in‐depth investigations of the structures of 2 and 3 in solution were carried out and a high‐resolution crystal structure of 2 was obtained. The two homologues of coenzyme B₁₂ ( 2 and 3 ) are suggested to function as covalent structural mimics of the hypothetical enzyme‐bound “activated” (that is, “stretched” or even homolytically cleaved) states of the B₁₂ cofactor. From crude molecular models, the crucial distances from the corrin‐bound cobalt center to the C5′ atom of the (homo)adenosine moieties in 2 and 3 were estimated to be about 3.0 and 4.4 Å, respectively. These values are roughly the same as those found in the two “activated” forms of coenzyme B₁₂ in the crystal structure of glutamate mutase. Indeed, in the crystal structure of 2 , the cobalt center was observed to be at a distance of 2.99 Å from the C5′ atom of the homoadenosine moiety and the latter was found to be present in the unusual syn conformation. In solution, the organometallic moieties of 2 and 3 were shown to be rather flexible and to be considerably more dynamic than the equivalent group in coenzyme B₁₂. The homoadenosine moiety of 2 was indicated to occur in both the syn and the anti conformations.     

TiO2‐assisted Photocatalytic Degradation of Vitamin B12 in Aqueous Solution:Influencing Factors,Kinetics, and Reaction Investigations

The photodegradation (λ=365 nm) of the biomolecule vitamin B₁₂, catalyzed by the photocatalyst TiO₂ nanoparticles (NPs), has been investigated in aqueous suspension. The photodegradation process of vitamin B₁₂ has been monitored by means of electronic absorption (Abs), Fourier‐transform infrared (FT‐IR), and resonance Raman (RR) spectroscopies, respectively. The results show that only under UV illumination in the presence of TiO₂ is there effective degradation, and the photocatalytic degradation of vitamin B₁₂ is strongly influenced by the amount of TiO₂ NPs, the pH, and the initial concentration of vitamin B₁₂. The photocatalytic reaction kinetics of vitamin B₁₂ conforms to a Langmuir‐Hinshelwood isotherm model. Changes involving the three structural units of the carbon‐metal bond C–Co, the organic corrin macrocycle combined with the benzimidazole nucleotide, and the inorganic CN in the vitamin B₁₂ molecule during the photocatalytic degradation are also discussed.     

Ultrasensitive Determination of Vitamin B12 Using Disposable Graphite Screen‐Printed Electrodes and Anodic Adsorptive Voltammetry

A facile, rapid and ultra‐sensitive method for the determination of vitamin B₁₂ (cyanocobalamin) at the sub‐nanomolar concentration range by using low‐cost, disposable graphite screen‐printed electrodes is described. The method is based on the cathodic preconcentration of square planar vitamin B_12s, as occurred due to the electro reduction of Co(III) center in vitamin B_12a to Co(I), at ?1.3 V versus Ag/AgCl/3 M KCl for 40 s. Then, an anodic square wave scan was applied and the height of the peak appeared at ca. ?0.73 V versus Ag/AgCl/3 M KCl, due to the oxidation of Co(I) to Co(II) in the adsorbed molecule, was related to the concentration of the vitamin B₁₂ in the sample. EDTA was found to serve as a key‐component of the electrolyte by eliminating the background signal caused by metal cations impurities contained in the electrolyte (0.1 M phosphate buffer in 0.1 M KCl, pH 3). It also blocks trace metals contained in real samples, thus eliminating their interference effect. The method was optimized to various working parameters and under the selected conditions the calibration curve was linear over the range 1×10^?10–8×10^?9 mol L^?1 vitamin B₁₂ (R²=0.994), while the limit of detection for a signal‐to‐noise ratio of 3 (7×10^?11 mol L^?1 vitamin B₁₂) is the lowest value of any reported in the literature for the electrochemical determination of vitamin B₁₂. The sensors were successfully applied to the determination of vitamin B₁₂ in pharmaceutical products.     

“Clickable” Vitamin B12 Derivative

A “clickable” vitamin B₁₂ derivative possessing the azide functionality at the 5′‐position was synthesized by means of a two‐step procedure on the gram scale. The reaction of cobalamin with mesyl chloride (MsCl) afforded the 5′‐OMs derivative, which was subsequently transformed to the desired 5′‐azide, the structure of which was confirmed using X‐ray analysis. It proved to be reactive in the azide–alkyne 1,3‐dipolar cycloaddition reaction to give substituted triazoles in high yields. A study of the reaction conditions and the scope of the process are reported.     

Solution,Crystal and in Silico Structures of the Organometallic Vitamin B12-Derivative Acetylcobalamin and of its Novel Rhodium-Analogue Acetylrhodibalamin

The natural vitamin B₁₂-derivatives are intriguing complexes of cobalt that entrap the metal within the strikingly skewed and ring-contracted corrin ligand. Here, we describe the synthesis of the Rh(III)-corrin acetylrhodibalamin ( AcRhbl ) from biotechnologically produced metal-free hydrogenobyric acid and analyze the effect of the replacement of the cobalt-center of the organometallic vitamin B₁₂-derivative acetylcobalamin ( AcCbl ) with its group-IX homologue rhodium, to give AcRhbl . The structures of AcCbl and AcRhbl were thoroughly analyzed in aqueous solution, in crystals and by in silico methods, in order to gain detailed insights into the structural adaptations to the two homologous metals. Indeed, the common, nucleotide-appended corrin-ligand in these two metal corrins features extensive structural similarity. Thus, the rhodium-corrin AcRhbl joins the small group of B₁₂-mimics classified as ‘antivitamins B₁₂’, isostructural metal analogues of the natural cobalt-corrins that hold significant potential in biological and biomedical applications as selective inhibitors of key cellular processes.     

Antivitamin B12 Inhibition of the Human B12-Processing Enzyme CblC: Crystal Structure of an Inactive Ternary Complex with Glutathione as the Cosubstrate

B₁₂ antivitamins are important and robust tools for investigating the biological roles of vitamin B₁₂. Here, the potential antivitamin B₁₂ 2,4-difluorophenylethynylcobalamin (F2PhEtyCbl) was prepared, and its 3D structure was studied in solution and in the crystal. Chemically inert F2PhEtyCbl resisted thermolysis of its Co−C bond at 100 °C, was stable in bright daylight, and also remained intact upon prolonged storage in aqueous solution at room temperature. It binds to the human B₁₂-processing enzyme CblC with high affinity (K_D=130 nm ) in the presence of the cosubstrate glutathione (GSH). F2PhEtyCbl withstood tailoring by CblC, and it also stabilized the ternary complex with GSH. The crystal structure of this inactivated assembly provides first insight into the binding interactions between an antivitamin B₁₂ and CblC, as well as into the organization of GSH and a base-off cobalamin in the active site of this enzyme.     

Antivitamins B12—Some Inaugural Milestones

The recently delineated structure- and reactivity-based concept of antivitamins B₁₂ has begun to bear fruit by the generation, and study, of a range of such B₁₂-dummies, either vitamin B₁₂-derived, or transition metal analogues that also represent potential antivitamins B₁₂ or specific B₁₂-antimetabolites. As reviewed here, this has opened up new research avenues in organometallic B₁₂-chemistry and bioinorganic coordination chemistry. Exploratory studies with antivitamins B₁₂ have, furthermore, revealed some of their potential, as pharmacologically interesting compounds, for inducing B₁₂-deficiency in a range of organisms, from hospital resistant bacteria to laboratory mice. The derived capacity of antivitamins B₁₂ to induce functional B₁₂-deficiency in mammalian cells and organs also suggest their valuable potential as growth inhibitors of cancerous human and animal cells.     

Probing Reversible Chemistry in Coenzyme B12‐Dependent Ethanolamine Ammonia Lyase with Kinetic Isotope Effects

Coenzyme B₁₂‐dependent enzymes such as ethanolamine ammonia lyase have remarkable catalytic power and some unique properties that enable detailed analysis of the reaction chemistry and associated dynamics. By selectively deuterating the substrate (ethanolamine) and/or the β‐carbon of the 5′‐deoxyadenosyl moiety of the intrinsic coenzyme B₁₂, it was possible to experimentally probe both the forward and reverse hydrogen atom transfers between the 5′‐deoxyadenosyl radical and substrate during single‐turnover stopped‐flow measurements. These data are interpreted within the context of a kinetic model where the 5′‐deoxyadenosyl radical intermediate may be quasi‐stable and rearrangement of the substrate radical is essentially irreversible. Global fitting of these data allows estimation of the intrinsic rate constants associated with Co?C homolysis and initial H‐abstraction steps. In contrast to previous stopped‐flow studies, the apparent kinetic isotope effects are found to be relatively small.     

7‐Decarboxymethyl‐cobyrinates: Vitamin B12‐Derivatives that Lack the c‐Side Chain

The synthesis of cobyrinic acid derivatives by reduction of dehydrocobyrinates is largely unexplored. It is, however, a rational path to B₁₂ analogues that lack specific substituents of the corrin moiety of natural B₁₂ derivatives. The partial syntheses of four epimeric 7‐decarboxymethyl‐cobyrinates is described, which is achieved by reduction of Δ7‐dehydro‐7‐de[carboxymethyl]‐cobyrinate with zinc or with the ‘prebiotic’ reducing agent formic acid. A direct and remarkably efficient route was found to 7‐decarboxymethyl‐cobyrinates, which are cobyrinic acid derivatives in which the c‐side chain at ring B of vitamin B₁₂ is missing. The structures of the hexamethyl‐7‐decarboxymethyl‐cobyrinates were characterized and the stereochemical and conformational properties at their newly saturated ring B were analyzed. The stereochemical outcome of the reduction was found to depend strongly on the reaction conditions. In 7‐decarboxymethyl‐cobyrinates, both peripheral carbon centres of ring B carry a hydrogen atom, and the characteristic quaternary carbon centre at C7 of the cobyrinic acid moiety of vitamin B₁₂ is lacking. The still highly substituted 7‐decarboxymethyl‐cobyrinates are readily dehydrogenated in the presence of dioxygen, furnishing 7‐de[carboxymethyl]‐Δ⁷‐dehydro‐cobyrinate as the common, unsaturated oxidation product. The noted stability of vitamin B₁₂ and of other Co^III‐cobyrinates in the presence of air is a consequence of their highly substituted corrin macrocycle, a finding of interest in the context of chemical rationalizations of the B₁₂ structure.     

New developments in the field of vitamin B12: enzymatic reactions dependent upon corrins and coenzyme B12.

Simple corrins such as vitamin B₁₂ and vitamin B₁₂ coenzyme catalyze a variety of unusual enzymatic reactions of which some are still without analogy in organic or organometallic chemistry. The mechanisms of these reactions are currently the subject of lively discussion. The present review focuses attention on new ideas about the mode of action of vitamin B₁₂ coenzymes in enzymatic reactions.     

Zinc Substitution of Cobalt in Vitamin B12: Zincobyric acid and Zincobalamin as Luminescent Structural B12‐Mimics

Replacing the central cobalt ion of vitamin B₁₂ by other metals has been a long‐held aspiration within the B₁₂‐field. Herein, we describe the synthesis from hydrogenobyric acid of zincobyric acid ( Znby ) and zincobalamin ( Znbl ), the Zn‐analogues of the natural cobalt‐corrins cobyric acid and vitamin B₁₂, respectively. The solution structures of Znby and Znbl were studied by NMR‐spectroscopy. Single crystals of Znby were produced, providing the first X‐ray crystallographic structure of a zinc corrin. The structures of Znby and of computationally generated Znbl were found to resemble the corresponding Co^II‐corrins, making such Zn‐corrins potentially useful for investigations of B₁₂‐dependent processes. The singlet excited state of Znby had a short life‐time, limited by rapid intersystem crossing to the triplet state. Znby allowed the unprecedented observation of a corrin triplet (E^T=190 kJ mol^?1) and was found to be an excellent photo‐sensitizer for ¹O₂ (Φ_Δ=0.70).     

Development of Disposable Carbon Nanofibers Electrodes Supported on Filters

The purpose of this work is to develop simple and reproducible ways to fabricate carbon nanofibers (CNFs) electrodes. Disposable CNFs electrodes were fabricated after filtering CNFs through polytetrafluoroethylene filters (PTFE), using directly as electrodes these filters with the CNFs film. The electrochemical behaviour of CNFs/PTFE electrodes were tested with ferrocenemethanol and tris(2,2′‐bypyridyl)dichlororuthenium(II) (Ru(bipy)₃²⁺). As a proof of concept of the analytical usefulness of the CNFs/PFTE electrodes, the study of vitamin B₁₂ was carried out. Multivariate calibrate has been successfully used to determine vitamin B₁₂ in a complex medium with vitamins B₁ and B₆ as interferents.     

Partial Synthesis of Coenzyme B12 from Cobyric Acid

Here we report the direct chemical synthesis of coenzyme B₁₂ (AdoCbl) from Co_β ‐5′‐deoxyadenosylcobyric acid (AdoCby) and the preparation of the latter from crystalline CN ,H₂O‐cobyric acid (CN ,H₂OC by). AdoCby is a suggested common key intermediate in the biosynthesis of AdoCbl and of other cobamides in microorganisms. AdoCby was thoroughly characterized by spectroscopic means, including homo‐nuclear and hetero‐nuclear NMR , as such data are not available in published work. AdoCbl was prepared from AdoCby in one‐step in over 85% yield, by covalent attachment in aqueous solution of the integral B₁₂‐nucleotide moiety using 1‐ethyl‐3‐(3‐dimethylaminopropyl)‐carbodiimide (EDC ·HC l) and N‐hydroxybenzotriazole (HOB t) as coupling reagents. By the same procedure crystalline vitamin B₁₂ (CNC bl) was also prepared in 92% yield from CN ,H₂OC by. Coordination of the B₁₂‐nucleotide base at the Co_α ‐face of AdoCby or of CN ,H₂OC by was indicated to assist in the efficient covalent coupling at the activated f‐side chain function to furnish the complete corrinoids AdoCbl and CNC bl.     

Experimental Study of Hydrogen Bonding Potentially Stabilizing the 5′‐Deoxyadenosyl Radical from Coenzyme B12

Coenzyme B₁₂ can assist radical enzymes that accomplish the vicinal interchange of a hydrogen atom with a functional group. It has been proposed that the Co? C bond homolysis of coenzyme B₁₂ to cob(II)alamin and the 5′‐deoxyadenosyl radical is aided by hydrogen bonding of the corrin C19? H to the 3′‐O of the ribose moiety of the incipient 5′‐deoxyadenosyl radical, which is stabilized by 30 kJ mol^?1 (B. Durbeej et al., Chem. Eur. J. 2009 , 15, 8578–8585). The diastereoisomers (R)‐ and (S)‐2,3‐dihydroxypropylcobalamin were used as models for coenzyme B₁₂. A downfield shift of the NMR signal for the C19? H proton was observed for the (R)‐isomer (δ=4.45 versus 4.01 ppm for the (S)‐isomer) and can be ascribed to an intramolecular hydrogen bond between the C19? H and the oxygen of CHOH. Crystal structures of (R)‐ and (S)‐2,3‐dihydroxypropylcobalamin showed C19? H???O distances of 3.214(7) Å (R‐isomer) and 3.281(11) Å (S‐isomer), which suggest weak hydrogen‐bond interactions (?ΔG<6 kJ mol^?1) between the CHOH of the dihydroxypropyl ligand and the C19? H. Exchange of the C19? H, which is dependent on the cobalt redox state, was investigated with cob(I)alamin, cob(II)alamin, and cob(III)alamin by using NMR spectroscopy to monitor the uptake of deuterium from deuterated water in the pH range 3–11. No exchange was found for any of the cobalt oxidation states. 3′,5′‐Dideoxyadenosylcobalamin, but not the 2′,5′‐isomer, was found to act as a coenzyme for glutamate mutase, with a 15‐fold lower k_cat/K_M than 5′‐deoxyadenosylcobalamin. This indicates that stabilization of the 5′‐deoxyadenosyl radical by a hydrogen bond that involves the C19? H and the 3′‐OH group of the cofactor is, at most, 7 kJ mol^?1 (?ΔG). Examination of the crystal structure of glutamate mutase revealed additional stabilizing factors: hydrogen bonds between both the 2′‐OH and 3′‐OH groups and glutamate 330. The actual strength of a hydrogen bond between the C19? H and the 3′‐O of the ribose moiety of the 5′‐deoxyadenosyl group is concluded not to exceed 6 kJ mol^?1 (?ΔG).     

Pulse Radiolysis and Ultra‐High‐Performance Liquid Chromatography/High‐Resolution Mass Spectrometry Studies on the Reactions of the Carbonate Radical with Vitamin B12 Derivatives

The reactions of the carbonate radical anion (CO₃ ^{. ?}) with vitamin B₁₂ derivatives were studied by pulse radiolysis. The carbonate radical anion directly oxidizes the metal center of cob(II)alamin quantitively to give hydroxycobalamin, with a bimolecular rate constant of 2.0×10⁹ M ^?1 s^?1. The reaction of CO₃ ^{. ?} with hydroxycobalamin proceeds in two steps. The second‐order rate constant for the first reaction is 4.3×10⁸ M ^?1 s^?1. The rate of the second reaction is independent of the hydroxycobalamin concentration and is approximately 3.0×10³ s^?1. Evidence for formation of corrinoid complexes differing from cobalamin by the abstraction of two or four hydrogen atoms from the corrin macrocycle and lactone ring formation has been obtained by ultra‐high‐performance liquid chromatography/high‐resolution mass spectrometry (UHPLC/HRMS). A mechanism is proposed in which abstraction of a hydrogen atom by CO₃ ^{. ?} from a carbon atom not involved in the π conjugation system of the corrin occurs in the first step, resulting in formation of a Co^III C‐centered radical that undergoes rapid intramolecular electron transfer to form the corresponding Co^II carbocation complex for about 50 % of these complexes. Subsequent competing pathways lead to formation of corrinoid complexes with two fewer hydrogen atoms and lactone derivatives of B₁₂. Our results demonstrate the potential of UHPLC combined with HRMS in the separation and identification of tetrapyrrole macrocycles with minor modifications from their parent molecule.     

Analysis of Levodopa in the Presence of Vitamin B6 Using Carbon Paste Electrode Modified with 1‐Butyl‐3 methylimidazolium Hexafluorophosphate and CuO Nanoparticles

A carbon paste electrode modified with 1‐butyl‐3‐methylimidazolium hexafluorophosphate ionic liquid (BMIPF6) and CuO nanoparticles (CuO/NPs) (CPE/BMIPF₆/CuO/NPs) was fabricated and used for square wave voltammetric analysis of levodopa in the presence of vitamin B₆. The elemental analysis, SEM and XRD methods were used for characterization of synthesized CuO nanoparticle. CPE/BMIPF₆/CuO/NPs exhibited high electrical conductivity toward the electro‐oxidation of levodopa at a pH=7.0 as best experimental condition. Using CPE/BMIPF₆/CuO/NPs the levodopa and vitamin B₆ peaks are separated and oxidized at potentials of 0.565 V and 0.835 V, respectively; hence levodopa can be detected in the presence of vitamin B₆. The electrochemical response shows a linear relationship from concentration of levodopa and vitamin B₆ in the ranges of 0.06‐1000 μM and 0.1‐700.0 μM, respectively. Finally, CPE/BMIPF₆/CuO/NPs were applied as high performance tool for determination of levodopa and vitamin B₆ in real samples.     

Vitamin B12 supported on graphene oxide: As a bio‐based catalyst for selective aerobic oxidation of alcohols

The environmental impact of chemical processes has now opened new windows of opportunity for bio‐based catalysts. In this paper a highly active bio‐based catalyst of vitamin B₁₂ supported on graphene oxide nanosheets is reported for the selective aerobic oxidation of alcohols to the corresponding carbonyl compounds. Operational simplicity, mild reaction conditions, high yield and selectivity, non‐hazardous nature, commercial availability and affordability are the main advantages of this novel catalytic system.     

Pseudocoenzyme B12 and Adenosyl‐Factor A: Electrochemical Synthesis and Spectroscopic Analysis of Two Natural B12 Coenzymes with Predominantly ‘Base‐off' Constitution

Pseudocoenzyme B₁₂ (=Coβ‐(5′‐deoxy‐5′‐adenosyl)‐(adenin‐7‐yl)cobamide; 1 ) and adenosyl‐factor A (=Coβ‐(5′‐deoxy‐5′‐adenosyl)‐(2‐methyladenin‐7‐yl)cobamide; 3 ) are two natural analogues of coenzyme B₁₂ (=adenosylcobalamin‐Coβ‐(5′‐deoxy‐5′‐adenosyl)‐(5,6‐dimethyl‐1H‐benzimidazolyl)cobamide; 2 ), where the Co‐coordinating 5,6‐dimethyl‐1H‐benzimidazole nucleotide base of 2 is replaced by the purine bases adenine and 2‐methyladenine. In contrast to 2 , which exists solely in the ‘base‐on' form, UV/VIS spectroscopy qualitatively indicates ‘base‐off' constitution for 1 and 3 in aqueous solution. (cf. the established ‘base‐off' form as unexpected binding mode of B₁₂ cofactors in several B₁₂‐dependent enzymes, such as in methionine synthase from Escherichia coli and in glutamate mutase from Clostridium cochlearium). In the present work, pseudocoenzyme B₁₂ ( 1 ) was synthesized in 85% yield by alkylation with 5′‐O‐tosyladenosine of (adenin‐7‐yl)cob(I)amide, which was produced electrochemically from pseudovitamin B₁₂ (Coβ‐cyano‐(adenin‐7‐yl)cobamide). Likewise, adenosyl‐factor A ( 3 ) was prepared in ca. 70% yield from factor A (=Coβ‐cyano‐(2‐methyladenin‐7‐yl)cobamide; 5 ). All the spectroscopic properties of 1 and 3 in aqueous solution indicated that these two Coβ‐(5′‐deoxy‐5′‐adenosyl)‐(adenin‐7‐yl)cobamides exist predominantly in a ‘base‐off' constitution, with minor but significant contributions of the ‘base‐on' form. From the UV/VIS spectra, the temperature‐dependent equilibrium constants of the ‘base‐off'/‘base‐on' reconstitution reaction were determined as K_on ( 1 )=0.30 and K_on ( 3 )=0.48 at 25°, corresponding to a contribution of the ‘base‐on' forms of 23% for 1 and of 32% for 3 .     

Design of a Vitamin B1‐Selective Electrode Based on an Ion‐Pair and Its Application to Pharmaceutical Analysis

Vitamin B₁‐selective electrodes with PVC membrane were developed that contain ion associates of vitamin B₁ with an inorganic anion, BiI₄^?, and an organic anion, brilliant yellow, as electrode‐active substances. The linearity ranges of the electrode function are 1.0×10^?5–1.0×10^?2 and 1.0×10^?4–1.0×10^?2 M, the electrode function slopes are 33.0±1.0 and 33.1±1.1 mV decade^?1, the detection limits are 5.5×10^?6 and 8.3×10^?5 M for BiI₄^? and brilliant yellow respectively. The working range of pH is 5–12. The efficiency of the use of electrodes for the vitamin B₁ content control in multivitamin pharmaceutical preparations was shown by direct potentiometry and potentiometric titration methods.     

Electrocrystallization of Tetra‐ and Hexa‐coordinated Silver(II) Compounds Based on 4,4′‐Dimethyl‐2,2′‐Bipyridine Ligand – Single Crystal Structures and Magnetic Studies

In this paper we report on the potential dependent electrocrystallization of [Ag(4,4′‐dimethyl‐2,2′‐bipyridine)₂(NO₃)₂] ( 1 ) and Ag(4,4′‐dimethyl‐2,2′‐bipyridine)(NO₃)₂ ( 2 ) from the same electrolytic bath. Thus it has been shown for the first time that the coordination number of silver ion to ligands can be tuned by the electrocrystallization potential. The single crystal structure analysis [ 1 : C2/c, a = 18.6308(15), b = 14.5708(12), c = 11.5867(10) Å, β = 126.5910(10)°, Z = 4, R = 3.9 %] [ 2 : P2₁/c, a = 8.5865(11) b = 11.0157(14) c = 16.4554(10) Å, β = 111.102(10), Z = 4 , R = 3.5 %] show divalent silver to be in an approximately square planar surrounding. Both complexes are paramagnetic following Curie's law with magnetic moments of 1.86 μ_B and 1.72 μ_B respectively.