Vitamin D4 in Mushrooms

Abstract

An unknown vitamin D compound was observed in the HPLC-UV chromatogram of edible mushrooms in the course of analyzing vitamin D2 as part of a food composition study and confirmed by liquid chromatography-mass spectrometry to be vitamin D4 (22-dihydroergocalciferol). Vitamin D4 was quantified by HPLC with UV detection, with vitamin [3H] itamin D3 as an internal standard. White button, crimini, portabella, enoki, shiitake, maitake, oyster, morel, chanterelle, and UV-treated portabella mushrooms were analyzed, as four composites each of a total of 71 samples from U.S. retail suppliers and producers. Vitamin D4 was present (>0.1 µg/100 g) in a total of 18 composites and in at least one composite of each mushroom type except white button. The level was highest in samples with known UV exposure: vitamin D enhanced portabella, and maitake mushrooms from one supplier (0.2–7.0 and 22.5–35.4 µg/100 g, respectively). Other mushrooms had detectable vitamin D4 in some but not all samples. In one composite of oyster mushrooms the vitamin D4 content was more than twice that of D2 (6.29 vs. 2.59 µg/100 g). Vitamin D4 exceeded 2 µg/100 g in the morel and chanterelle mushroom samples that contained D4, but was undetectable in two morel samples. The vitamin D4 precursor 22,23-dihydroergosterol was found in all composites (4.49–16.5 mg/100 g). Vitamin D4 should be expected to occur in mushrooms exposed to UV light, such as commercially produced vitamin D enhanced products, wild grown mushrooms or other mushrooms receiving incidental exposure. Because vitamin D4 coeluted with D3 in the routine HPLC analysis of vitamin D2 and an alternate mobile phase was necessary for resolution, researchers analyzing vitamin D2 in mushrooms and using D3 as an internal standard should verify that the system will resolve vitamins D3 and D4.

Impact on Vitamin D2, Vitamin D4 and Agaritine in Agaricus bisporus Mushrooms after Artificial and Natural Solar UV Light Exposure

“…The moisture contents found in the present study are in agreement with published data [11], however a limitation of the study was the use of the freeze-drying factor for determination of moisture content as this method does not account for possible residual moisture, which could be up to 6 % [34].…”

“…Figure 3 illustrates vitamin D 4 content as a function of the vitamin D 2 content in sliced mushrooms exposed to irradiation from UV-B tubes. We detected no vitamin D 4 in the non-exposed control samples, concurring with analyses of button mushrooms not detecting vitamin D 4 in 10 of 12 samples [11]. Those researchers observed vitamin D 4 in 41 % of non-exposed mushroom samples of different species, with higher contents in wild-grown mushrooms.…”

“…The solution was saponified, liquid-liquid extracted, and clean-up through a silica solid phase extraction and a preparative normal phase HPLC for collection of the vitamin D fraction. No commercial standard is available for vitamin D 4 , therefore the identification of vitamin D 4 was [16] b Distance of 75 cm between the tubes and the samples and an irradiation intensity of 0.69 ± 0.04 mW/cm 2 measured at 18 different positions c Distance of 137 cm between the tubes and the samples and an irradiation intensity of 0.15 mW/cm 2 based on information provided by Philips et al [11] identifying vitamin D 4 in UV-exposed mushrooms. Following evaporation and dilution in mobile phase [acetonitrile:methanol (80: 20)] for the analytical HPLC-system (VYDAC201TP54, 5 μm, 250 × 4.6 mm, Grace, MD) the vitamin D 2 , D 3 , and D 4 were separated having retention times at 19.6, 20.0, and 23.4 min, respectively.…”

“…There is published evidence that the vitamin D 4 precursor sterol is detectable in all mushrooms (varying widely among species) and comparable to vitamin D 2 precursor convertible to vitamin D 4 via a photochemical reaction [11]. Figure 3 illustrates vitamin D 4 content as a function of the vitamin D 2 content in sliced mushrooms exposed to irradiation from UV-B tubes.…”

“…They were the first to assess the agaritine content, finding that it did not change as a result of exposure to sunlight or artificial UV-B. In addition, Phillips et al [11] demonstrated the omnipresence of the vitamin D 4 precursor 22,23-dihydroergosterol in mushrooms and hypothesised the occurrence of vitamin D 4 in UV-exposed mushrooms [11]. Agaritine [N-(gamma-Lglutamyl)-4-hydroxymethylphenylhydrazine] is an aromatic hydrazine found in mushrooms of the genus Agaricus.…”