Analysis of Salazinic, Norstictic, and Usnic Acids in Xanthoparmelia chlorochroa by Ultra-Performance Liquid Chromatography/Tandem Mass Spectrometry

Abstract

The lichen species Xanthoparmelia chlorochroa is toxic when consumed by domestic sheep, cattle, and Rocky Mountain elk. Clinical signs exhibited by poisoned animals include red urine, ataxia, and muscular weakness that rapidly progresses to recumbency. Elk are unable to recover once becoming recumbent; however, most affected cattle can recover if offered suitable feed shortly following the onset of signs. At present, the pathogenesis and specific toxin(s) are unknown. As part of an effort to elucidate the proximate toxin, a method using ultra-performance LC coupled to MS/MS with negative-ion electrospray ionization has been developed to compare salazinic, norstictic, and usnic acid concentrations in X. chlorochroa collected from locales associated with lichen poisonings. Compounds were extracted from lichen samples with acetone and sonication. The stationary phase was a Waters Acquity UPLCTM BEH C18 (50 2.1 mm; 1.7 m particle size) column. The mobile phase consisted of an acetonitrilewater gradient. The precision of the method was confirmed by an SD below 0.4 (n = 9) for triplicate samples. LOD values were 200, 100, and 50 ng/mL for salazinic, norstictic, and usnic acids, respectively.

Methodological Aspects of Green Extraction of Usnic Acid Using Natural Deep Eutectic Solvents

“…The four independent variables, including the camphor:thymol (X1) molar ratio, the Although the solubility of UA in acetone (see Table 1) allowed exhaustive extraction from the raw material (1.45 mg of UA from 100 mg of raw material with a UA solubility of about 9.55 mg/1.5 mL of acetone), the extraction yield was relatively low, and the subsequent extraction steps increased the yield only slightly. This phenomenon showed that, although acetone is the most common solvent used for the extraction of lichen metabolites [5,10,[14][15][16][17][18][19][20], this extractant did not provide exhaustive extraction of UA from C. uncialis.…”

“…Usnic acid (UA) is a phenolic compound with a dibenzofuran structure occurring in the form of two enantiomers ( Figure 1 ) [ 1 ] in several lichen genera ( Alectoria , Cladonia , Evernia , Lecanora , Parmelia , Ramalina , Usnea , and Xanthoparmelia ) as a yellow cortical pigment [ 2 , 3 , 4 , 5 , 6 ]. Nowadays, it is the most extensively investigated lichen secondary metabolite.…”

“…The values of the LogP coefficient (2.88) [ 12 ] and topological polar surface area (118 Å 2 ) [ 13 ] evidence the lipophilic nature of this compound, which largely determines its poor solubility in polar solvents. For this reason, UA is extracted from lichens using solvents such as acetone [ 5 , 10 , 14 , 15 , 16 , 17 , 18 , 19 , 20 ], dichloromethane [ 21 , 22 , 23 , 24 ], petroleum ether [ 3 , 4 , 25 ], n-hexane [ 26 ], and heptane [ 27 ]. Methanol and ethanol are also used as solvents, although much less frequently [ 4 , 28 , 29 ], as their extraction yields are lower than those of non-polar solvents.…”

Morphological, Secondary Metabolite and ITS (rDNA) Variability within Usnic Acid‐Containing Lichen Thalli of Xanthoparmelia Explored at the Local Scale of Rock Outcrop in W‐Alps

“…[23] Some of these show antioxidant [24] and cancer-chemopreventive activity, [25] as well as UV screening capacity, [26] defence functions against herbivores [27] and toxic effects to domestic and wild mammals. [28] Crude extracts from X. chlorochroa (TUCKERMAN) HALE exhibited antiproliferative effects against lymphoma (Rajii) cells. [19] Although Xanthoparmelia lichen thalli may thus appear a promising source of natural products, they may also pose difficulties and limitations for the collection of homogeneous materials in bulk quantities to test and exploit LSMs [22] : the high LSMs availability lies within a complex framework of variability, which is still widely unresolved.…”

“…Lichens of the world‐wide distributed genus Xanthoparmelia ( Vainio ) Hale are known to synthesize more than 90 LSMs . Some of these show antioxidant and cancer‐chemopreventive activity, as well as UV screening capacity, defence functions against herbivores and toxic effects to domestic and wild mammals . Crude extracts from X .…”