J Nutr Health.  2018 Aug;51(4):275-286. 10.4163/jnh.2018.51.4.275.

Metabolites profiling and hypolipidemic/hypocholesterolemic effects of persimmon (Diosyros kaki Thumb.) by different processing procedures: in vitro and in vivo studies

• Park SY
• Oh EK
• Lim Y
• Shin JY
• Jung HA
• Park SY
• Lee JH
• Choe JS
• Kwon O

• KMID: 2419498
• DOI: http://doi.org/10.4163/jnh.2018.51.4.275

Abstract

PURPOSE
Our previous study demonstrated that persimmon (Diospyros kaki Thumb.) at different stages of ripening provided different protective effects against high-fat/cholesterol diet (HFD)-induced dyslipidemia in rats. In this study, we compared the metabolites profile and gene expressions related to triglyceride (TG)/cholesterol metabolism in vitro and in vivo after treating with persimmon water extracts (PWE) or tannin-enriched persimmon concentrate (TEP).
METHODS
Primary and secondary metabolites in test materials were determined by GC-TOF/MS, UHPLC-LTQ-ESI-IT-MS/MS, and UPLC-Q-TOF-MS. The expression of genes related to TG and cholesterol metabolism were determined by RT-PCR both in HepG2 cells stimulated by oleic acid/palmitic acid and in liver tissues obtained from Wistar rats fed with HFD and PWE at 0, 150, 300, and 600 mg/d (experiment I) or TEP at 0, 7, 14, and 28 mg/d (experiment II) by oral gavage for 9 weeks.
RESULTS
PLS-DA analysis and heatmap analysis demonstrated significantly differential profiling of metabolites of PWE and TEP according to processing of persimmon powder. In vitro, TEP showed similar hypolipidemic effects as PWE, but significantly enhanced hypocholesterolemic effects compared to PWE in sterol regulatory element-binding protein 2 (SREBP2), HMG-CoA reductase (HMGCR), proprotein convertase subtilisin/kexin type 9 (PCSK9), cholesterol 7α-hydroxylase (CYP7A1), and low density lipoprotein receptor (LDLR) gene expression. Consistently, TEP and PWE showed similar hypolipidemic capacity in vivo, but significantly enhanced hypocholesterolemic capacity in terms of SREBP2, HMGCR, and bile salt export pump (BSEP) gene expression.
CONCLUSION
These results suggest that column extraction after hot water extraction may be a good strategy to enhance tannins and long-chain fatty acid amides, which might cause stimulation of hypocholesterolemic actions through downregulation of cholesterol biosynthesis gene expression and upregulation of LDL receptor gene expression.

Keyword

Diospyros kaki Thumb.; metabolites profiling; Hep G2 cells; diet-induced hyperlipidemic rats

MeSH Terms

Amides
Animals
Bile
Cholesterol
Diet
Diospyros*
Down-Regulation
Dyslipidemias
Gene Expression
Hep G2 Cells
In Vitro Techniques*
Liver
Metabolism
Oxidoreductases
Proprotein Convertases
Rats
Rats, Wistar
Receptors, LDL
Tannins
Triglycerides
Up-Regulation
Water
Amides
Cholesterol
Oxidoreductases
Proprotein Convertases
Receptors, LDL
Tannins
Water

Figure

• 

  Fig. 1 Partial lest-squares discrimination analysis (PLS-DA) score plots using the GC-TOF-MS (A) and UHPLC-LTQ-ESI-IT-MS/MS/UPLC-Q-TOF-MS (B) data sets and heat map of specific metabolites in PDP, PWE, and TEP (C). PDP, persimmon dried powder; PWE, persimmon water extract; TEP, tannin-enriched persimmon concentrate. R2X and R2Y were the cumulative modeled variation in X and Y matrix, respectively. Q2 were the cumulative predicted variation in Y matrix.

• 

  Fig. 2 Effects of PWE and TEP on intracellular lipid accumulation (A and B, respectively) and FAS gene expression (C) in HepG2 cells stimulated by free fatty acids (0.66 mM oleic acid + 0.33 mM palmitic acid). PWE, persimmon water extract; TEP, tannin-enriched persimmon concentrate. HepG2 cells were grown in serum-free medium overnight and incubated in 1mM OA/PA-BSA complex in the absence or presence of PWE (A) or TEP (B) for an additional 24h. Values are means from three independent experiments conducted with triplicate treatments, with standard error represented by vertical bars. *Significantly different at p < 0.05 by Dunnett's test compared with FFA loading.

• 

  Fig. 3 Effects of PWE and TEP on cholesterol synthesis (A) and cholesterol metabolism (B) in HepG2 cells stimulated by free fatty acids (0.66 mM oleic acid + 0.33 mM palmitic acid). PWE, persimmon water extract; TEP, tannin-enriched persimmon concentrate. HepG2 cells were grown in serum-free medium overnight and incubated in 1mM OA/PA-BSA complex in the absence or presence of PWE (A) or TEP (B) for an additional 24h. Values are means from three independent experiments conducted with triplicate treatments, with standard error represented by vertical bars. *Significantly different at p < 0.05 by Dunnett's test compared with FFA loading.

• 

  Fig. 4 Effects of PWE (A) and TEP (B) on hepatic lipogenesis (SREBP1c and FAS) in rats fed a high-fat/cholesterol diet. PWE, persimmon water extract; TEP, tannin-enriched persimmon concentrate, HF, high-fat/cholesterol control; HL, HF + low-dose PWE/TEP; HM, HF + middle-dose PWE/TEP; HH, HF + high-dose PWE/TEP. Values are expressed as mean ± SE (n = 10 for each group). *Significantly different at p < 0.05 by Dunnett's test compared with HF group.

• 

  Fig. 5 Effects of PWE (A) and TEP (B) on hepatic cholesterol metabolism (SREBP2, LDLR, HMGCR and CYP7A1, BSEP) in rats fed a high-fat/cholesterol diet. PWE, persimmon water extract; TEP, tannin-enriched persimmon concentrate, HF, high-fat/cholesterol control; HL, HF + low-dose PWE/TEP; HM, HF + middle-dose PWE/TEP; HH, HF + high-dose PWE/TEP. Values are expressed as mean ± SE (n = 10 for each group). *Significantly different at p < 0.05 by Dunnett's test compared with HF group.

• 

  Fig. 6 Overview diagram of research scope and flow

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