Lowering of lysophosphatidylcholines in ovariectomized rats by Curcuma comosa

Abstract

Decline of ovarian function in menopausal women increases metabolic disease risk. Curcuma comosa extract and its major compound, (3R)-1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol (DPHD), improved estrogen-deficient ovariectomized (OVX) rat metabolic disturbances. However, information on their effects on metabolites is limited. Here, we investigated the impacts of C. comosa ethanol extract and DPHD on 12-week-old OVX rat metabolic disturbances, emphasizing the less hydrophobic metabolites. Metabolomics analysis of OVX rat serum showed a marked increase compared to sham-operated rat (SHAM) in levels of lysophosphatidylcholines (lysoPCs), particularly lysoPC (18:0) and lysoPC (16:0), and of arachidonic acid (AA), metabolites associated with inflammation. OVX rat elevated lysoPCs and AA levels reverted to SHAM levels following treatments with C. comosa ethanol extract and DPHD. Overall, our studies demonstrate the effect of C. comosa extract in ameliorating the metabolic disturbances caused by ovariectomy, and the elevated levels of bioactive lipid metabolites, lysoPCs and AA, may serve as potential biomarkers of menopausal metabolic disturbances.

Fig 1

Schematic diagram of (A) experimental procedures and (B) metabolomics analysis.

Fig 2. Multivariate data analysis of untargeted metabolomics profiles.

(A) Principal component analysis. (B) Orthogonal projection to latent structure with discriminant analysis. Data were obtained from LC–MS-based untargeted metabolomics analysis of serum samples of sham-operated (SHAM) (green circles) and bilateral ovariectomized (OVX) (blue squares) rats at week 12 post-treatment (n = 7 per group). PC, principal component.

Fig 3. Identification of lysophosphatidylcholines (lysoPCs) as OVX-associated metabolites.

(A) S-loading plot of serum metabolic profiles in orthogonal projection to latent structure with discriminant analysis (OPLS-DA). Each square represents a metabolite feature. The upper right and lower left quadrant contains metabolite features with increased and decreased levels in OVX compared to SHAM samples, respectively. Heat scale (Density) indicates Z-score, ranging from highest (red) to lowest (blue). p(corr)[1], reliability; p[1], magnitude. (B) LC–MS/MS spectrum of protonated lysoPC (18:0) fragment ions at 40 eV. Ions of m/z 86.0967, 104.1066, and 184.0735 arose from the phosphocholine head group, and ions of m/z 341.3036 and 506.3579 arose from the loss of water or phosphocholine from the protonated lysoPC molecular ion of m/z 524.3715 (blue diamond).

Fig 4. Representative GC–MS spectrum of trimethylsilyl-derivatized cholesterol.

Red, MS spectrum of trimethylsilyl-derivatized cholesterol from serum of ovariectomized rat; blue, spectrum from GC–MS NIST library.

Fig 5. Heat map of rat serum metabolite levels.

Rat serum metabolites were quantified by LC–and GC–MS-based metabolomics analysis. Each square represents serum data from each sham-operated (SHAM), bilateral ovariectomized (OVX), OVX treated with diarylheptanoid (3R)-1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol (DPHD), and OVX treated with Curcuma comosa ethanol extract (EXT) rat (n = 7 per group). Z-score scale ranges from highest (red) to lowest (green) abundance level.

Fig 6. Relative levels of potential serum biomarkers of metabolic changes arising from ovariectomy.

Rat serum metabolites were quantified by LC–and GC–MS-based metabolomics analysis. Results are shown as mean ± SEM (n = 7) metabolite peak area (concentration) relative to sham-operated rat (SHAM) of bilateral ovariectomized (OVX), OVX treated with diarylheptanoid (3R)-1,7-diphenyl-(4E,6E)-4,6-heptadien-3-ol (DPHD), and OVX treated with Curcuma comosa ethanol extract (EXT) rat. *p-value <0.05, **p-value <0.01 compared to SHAM. ^†p-value <0.05, ^†† p-value <0.01 compared to OVX.