Effects of Chinese Dietary Pattern of Fat Content, n-6/n-3 Polyunsaturated Fatty Acid Ratio, and Cholesterol Content on Lipid Profile in Rats.

This study aims to investigate the effect of Chinese diet pattern of fat content (30% or 36.06%), n-6/n-3 polyunsaturated fatty acid (PUFA) ratio (5 : 1 or 9 : 1), and cholesterol content (0.04 or 0.057 g/kg total diet) on lipid profile using a rat model. Results showed that rats' body weights (BWs) were controlled by the simultaneous intakes of cholesterol level of 0.04 g/kg total diet and n-6/n-3 ratio of 5 : 1. In addition, under high-fat diet, increased cholesterol feeding led to increased total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) levels and decreased triacylglycerols (TG) in rats' plasma. However, high density lipoprotein cholesterol (HDL-C) level and the ratios of HDL-C/LDL-C and HDL-C/TC in rats' plasma increased in response to simultaneous intakes of low n-6/n-3 ratio (5 : 1) and cholesterol (0.04 g/kg total diet) even under high-fat diet. Moreover, as the n-6/n-3 PUFA ratio in the diet decreased, the proportion of n-3 PUFAs increased in plasma, liver, and muscle and resulted in the decrease of n-6/n-3 PUFA ratio.

1. Introduction

The amount and type of dietary fats largely influence plasma lipid content and play an important role in diseases prevention and development [1, 2]. In recent decade years, the number of obese and/or overweight people in China was up to 2,600 million and the data are increasing, which was largely associated with the unhealthy diet of high fat and cholesterol [3]. Data from National Nutrition and Health Survey in China showed that the average intake of fat was 76.2 g/60 kg·body weight (BW)·d in the whole country (30% total calories) and 85.6 g/60 kg·BW·d in the urban areas (36.06% total calories) [4]. And the consumption ratio of n-6/n-3 was 9 : 1 in both the whole country and the urban areas [5]. Moreover, the average cholesterol consumption in the whole country and the urban areas was 0.04 and 0.057 g/kg total diet, respectively [4, 5]. It is obvious that the Chinese dietary pattern of fat and cholesterol has surpassed the recommended level and the ratio of n-6/n-3 almost reached the upper level of FAO/WHO standards [6]. Besides, it was found that the increased Chinese dietary pattern of fat and cholesterol was positively associated with the increased morbidity and mortality of cardiovascular disease, with R2 value of 0.98 and 1.00, respectively [5]. However, to the best of our knowledge, little has been reported regarding the influence of Chinese dietary pattern of fat and cholesterol on lipid metabolism.

Currently, there are many studies, using rat models, investigating the effects of different fatty acid ratios with or without cholesterol treatment on rats' lipid metabolism. For example, Parrish et al. [7] found that there were attenuated perirenal and epididymal fat accumulation and reduction in adipocyte size in rats fed with a high-fat diet consisting of fish oil, compared to those fed with lard. Wong et al. [8] and Balasubramaniam et al. [9] reported that, in rats, dietary long-chain n-3 polyunsaturated fatty acid (PUFA) could reduce both triacylglycerols (TG) and cholesterol plasma concentrations. Yang et al. [10] found that comparing with rat group fed with n-6/n-3 ratio of 20 : 1, rat groups with the ratios of 1 : 1 and 5 : 1 had significantly decreased serum levels of total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), and proinflammatory cytokines. Besides, Huang et al. [11] reported that the cholesterol-added diet (1%) did not impair the response to long-chain n-3 PUFA but increased the plasma cholesterol content and the liver weight. Moreover, Lu and Wu [12] found that added 1% cholesterol to diets led to a reduction of plasma TG and high density lipoprotein (HDL) levels and an increase of liver TG and cholesterol levels.

Therefore, the aim of this study was to investigate the influence of different fat intake, n-6/n-3 ratio, and cholesterol on lipids metabolism using a rat model according to the Chinese diet pattern. BWs, plasma lipid levels, and fatty acid compositions in rats' plasma and tissues were determined and analyzed.

2. Methods and Materials

2.1. Materials and Chemicals

A total of 72 young male Sprague-Dawley rats (two or three months, 270 ± 33 g at the beginning of the experiment) were purchased from Experimental Animal Center of Jiangxi Medical College, China (approval number: 02196-02). They were randomly divided into 6 groups with 12 rats per group (Table 1) according to the different consumption of fat energy (30 or 36.06%), n-6/n-3 ratio (5 : 1 or 9 : 1), and cholesterol (0.04 or 0.057 g/kg total diet) and then were housed in the individual cages at a constant temperature of 22 ± 1°C with a normal 12 : 12 hours of light dark cycles. The six groups were Experimental Groups 1, 2, 3, and 4 (EG 1, 2, 3, and 4) and Control Groups 1 and 2 (CG 1 and 2). Except for different contents of palm oil and cholesterol in different groups, the compositions of other nutrients in the diet given to rats were identical (Table 2). Besides, rats in each group was intragastric gavage administered with blended oil of 0.8 mL/100 g·BW·d (EG 1, 2, 3, and 4) or 0.6 mL/100 g·BW·d (CG 1 and 2), so as to maintain the dietary consumption of fat energy and n-6/n-3 ratio in different group. The physical blended oil administered in each group consists of different proportions of camellia oil, sunflower oil, and linseed oil (fatty acid compositions shown in Table 3), and their formula in each group was shown in Table 4. All procedures involving animals were approved by the Ethics Committee, Nanchang University, and conducted in accordance with the guidelines of China for the care and use of laboratory animals.

Table 1

The grouping of rats according to the diet of Chinese people.

	bThe energy proportion of fat (%)	cThe dose for oral administration of the blended oil in rats (mL/100 g·BW·d)	dThe cholesterol level (g/kg total diet)	n-6/n-3
aEG 1	36.06	0.8	0.04	9 : 1
EG 2	36.06	0.8	0.057	9 : 1
EG 3	36.06	0.8	0.04	5 : 1
EG 4	36.06	0.8	0.057	5 : 1
CG 1	30	0.6	0.04	9 : 1
CG 2	30	0.6	0.057	9 : 1

aEG was experimental group and CG was control group. bThe energy of fat consumed in the urban areas (36.06%) and in the whole country (30%), respectively. cAverage intake of blended oils in rats converted from Chinese dietary consumption per day in the urban areas (40 g/60 kg·BW·d) and in the whole country (30 g/60 kg·BW·d), respectively. dThe cholesterol proportion of total diet in the whole country (0.04 g/kg total diet) and the urban areas (0.057 g/kg total diet), respectively.

Table 2

The components and nutrition compositions of basal chow and the amount of palm oil and cholesterol of each group.

Basal components (g/kg)	Groups
	EG1	EG2	EG3	EG4	CG1	CG2
Corn flour	480	480	480	480	480	480
Soybean flour	210	210	210	210	210	210
Wheat bran	170	170	170	170	170	170
Wheat flour	50	50	50	50	50	50
Fish meal	50	50	50	50	50	50
CaH2PO4	15	15	15	15	15	15
Limestone power	15	15	15	15	15	15
Salt	5	5	5	5	5	5
Bean oil	6	6	6	6	6	6
Nutrient compositions (%)
Carbohydrate	52.58
Fat	3.50
Protein	21.2
Moisture	8.80
Ash	6.78
Fiber	2.80
Additional components
Palm oil	90	90	86	86	61	61
Cholesterol (mg/kg)	40	57	40	57	40	57

EG was experimental group and CG was control group. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

Table 3

Main fatty acid compositions (% area) of four oils used.

Fatty acids	Palm oil	Camellia oil	Sunflower oil	Linseed oil
14:0	1.19	0.04	0.06	0.05
16:0	60.28	8.79	6.34	5.19
9c16:1	0.11	0.09	0.06	0.06
18:0	5.66	2.29	5.67	3.52
9t18:1	0.18	2.33	1.71	1.55
9c18:1 OA	23.98	76.46	19.12	20.35
11c18:1	1.41	ND	1.41	1.35
9c12c18:2n-6 LA	5.51	6.88	58.66	14.31
20:0	0.36	0.05	0.30	0.23
18:3n-6	ND	ND	ND	0.83
5c20:1	ND	0.02	ND	0.17
11c20:1	0.07	0.37	0.11	5.73
18:3n-3 ALA	0.21	0.28	0.21	34.65
21:0	ND	0.02	0.19	ND
22:0	0.05	0.01	0.56	0.07
22:1n-9	ND	0.03	ND	0.31
23:0	0.03	0.01	ND	ND
24:0	0.06	0.03	0.11	ND
TSFA	67.73	11.31	13.23	9.06
TMUFA	25.57	76.95	20.74	27.66
TPUFA	5.73	7.26	58.87	50.28
n-6/n-3	26.29	24.64	279.33	0.44

OA, oleic acid; LA, linoleic acid; ALA, alpha-linolenic acid; TSFAs, total saturated fatty acids, the sum of (C14:0 + C16:0 + C18:0 + C20:0 + C21:0 + C22:0 + C23:0 + C24:0 + ⋯); TMUFAs, total monounsaturated fatty acids, the sum of (C16:1 + C18:1 + C20:1 + C22:1 + ⋯); TPUFAs, total polyunsaturated fatty acids, the sum of (C18:2n-6 + C18:3n-6 + C18:3n-3 + ⋯). ND, not detected. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

Table 4

The formulation design of blended oil intragastric gavage administered in each group.

Oil (g/100 g)	EG1/EG2	EG3/EG4	CG1/CG2
Camellia oil	10	18.6	10.2
Sunflower oil	74	53.5	73.5
Linseed oil	16	27.9	16.3
Total	100	100	100
Final fatty acid ratios administered to rats considering the oils in the chow
S : M : P	1 : 1.22 : 1	1 : 1.19 : 1	1 : 1.21 : 1.03
n-6/n-3	9.12	5.04	9.07

EG was experimental group and CG was control group. S: total saturated fatty acids; M: total monounsaturated fatty acids; TPUFA: total polyunsaturated fatty acids. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

2.2. Animal Treatment and Sampling

Before treatment, blood samples (about 0.5–1.0 mL) were collected from rats' tail vein to measure the biochemical indices, such as TC, TG, LDL-C, and HDL-C. TC and TG were measured enzymatically on the automatic biochemistry analyzer (Beckman, CX4) because of their methodology, calibrators, and controls [13]. The very low density lipoprotein cholesterol (VLDL-C) and LDL-C were precipitated using dextran sulfate and magnesium sulfate before HDL-C was measured enzymatically on the same instrument. LDL-C was calculated from these values (TC, HDL-C, and TG) using the formula of Friedewald et al. [14] when the TG level was <4.52 mmol/L.

After 7-day basal chow, there was no difference in BWs or serum lipid profile among the six groups. Then, each group was maintained on the experimental diet for 60 days. Rats had free access to water. BWs were monitored weekly. After 60 days' treatment, rats were fasted overnight (about 12 h, with water ad libitum) to reduce the feeding impact and then sacrificed under diethyl ether anesthesia. Blood samples were collected by cardiac puncture into clean test tubes containing 1% heparin sodium solution (w/w), centrifuged (4°C, 2500 r/min for 15 min) to get the plasma fraction, and then promptly stored at −80°C until analysis. The main organs and tissues (brain, heart, liver, and muscle) were removed and immediately rinsed in phosphate-buffered saline twice and then stored at −80°C until analysis.

2.3. Measurement of Biochemical Indices

Plasma lipid levels of TG, TC, LDL-C, and HDL-C after treatment were determined by automatic biochemistry analyzer (CX4, Beckman) as described above.

2.4. Fatty Acid Composition Analysis in Blood and Tissues

Total lipids were extracted with chloroform/methanol (v/v, 1 : 1) from the liver, brain, heart, muscle, and plasma according to the method of Folch et al. [15]. The obtained lipids were converted to fatty acid methyl esters (FAMEs) following the method of Lei et al. [16] and then analyzed by gas chromatography (model 6890N, Agilent Technologies, USA) equipped with an autoinjector, a fused-silica capillary column (CP-Sil 88, 100 m × 0.25 mm × 0.2 μm i.d.), and a flame ionization detector (FID) and operated on in splitless mode. The injector and detector temperatures were maintained at 250 and 260°C, respectively. The column temperature was held at 45°C for 3 min and then programmed to 175°C for 27 min at the rate of 13°C/min. The temperature was then further increased to 215°C at the rate of 4°C/min and held for 35 min. The oven temperature program was sustained for 86 min in total. The carrier gas was nitrogen, and the total gas flow rate was 52 mL/min. The injecting sample volume was 1 μL. Analysis of all peaks was accomplished by comparison of their retention times with FAME standards.

2.5. Statistical Analysis

Data were presented as mean ± SD and carried out using the Kruskal-Wallis one-way analysis by Statistical Package for the Social Sciences (SPSS) to compare differences between groups. P values less than 0.05 (P < 0.05) were considered significant.

3. Results

3.1. BWs

Under different diet of fat, n-6/n-3 ratio, and cholesterol, the change of BWs among six groups showed difference from the fourth week (Table 5). Compared with CG2 fed with fat energy of 30%, average BW in EG2 fed with higher fat energy of 36.06% was significantly increased (P < 0.05). Besides, with the same fat energy of 36.06% and n-6/n-3 ratio of 9 : 1, but different cholesterol level of 0.04 or 0.057 g/kg total diet, there was no significant difference (P > 0.05) of BWs in EG1 and EG2. Also, with the same fat energy of 30% and n-6/n-3 of 9 : 1, but different cholesterol level of 0.04 or 0.057 g/kg total diet, there was no significant difference (P > 0.05) of BWs in CG1 and CG2. However, with the same fat energy of 36.06% and n-6/n-3 of 5 : 1, the average BW in EG3 feeding cholesterol of 0.04 g/kg total diet was significantly lower than that in EG4 feeding cholesterol of 0.057 g/kg total diet (P < 0.05). Rats in EG4 fed with low n-6/n-3 ratio (5 : 1) but high cholesterol (0.057 g/kg total diet) showed no significant difference in BWs compared to those in EG1 feeding high n-6/n-3 ratio (9 : 1) but low cholesterol (0.04 g/kg total diet). Interestingly, rats in EG3 taking low n-6/n-3 ratio (5 : 1) and cholesterol (0.04 g/kg total diet) showed significantly decreased BWs compared to those in EG2 taking high n-6/n-3 ratio (9 : 1) and cholesterol (0.057 g/kg total diet). All these results suggested that rats' BW increased as the increase of fat energy, whereas the increase slowed down by reducing the intake of cholesterol under low intake of n-6/n-3 ratio.

Table 5

The body weights (g) of rats in weeks.

The week	EG1	EG2	EG3	EG4	CG1	CG2
0th	277 ± 26	277 ± 31	277 ± 40	277 ± 40	278 ± 32	277 ± 32
1st	307 ± 20	313 ± 25	313 ± 27	318 ± 32	316 ± 32	306 ± 19
2nd	349 ± 28	351 ± 23	349 ± 28	348 ± 32	343 ± 35	330 ± 19
3rd	385 ± 34	386 ± 21	361 ± 32	373 ± 38	370 ± 38	362 ± 22
4th	412 ± 30c	410 ± 22c	368 ± 31a	401 ± 30bc	390 ± 29ac	380 ± 27ab
5th	426 ± 32c	429 ± 22c	374 ± 29a	423 ± 33c	413 ± 30bc	391 ± 25ab
6th	447 ± 27c	435 ± 21c	378 ± 25a	444 ± 26c	424 ± 29bc	393 ± 30ab
7th	458 ± 32c	450 ± 25c	378 ± 26a	456 ± 31c	439 ± 31bc	412 ± 29ab
8th	466 ± 30c	463 ± 26c	375 ± 28a	460 ± 31c	452 ± 29bc	423 ± 27b
9th	481 ± 25c	474 ± 28c	376 ± 26a	465 ± 28c	462 ± 30bc	431 ± 29bc

Values presented as means (n = 12) ± SD. Mean values within each row followed by different letters (a, b, c) are significantly (P < 0.05) different. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

3.2. The Plasma Lipid Levels

As shown in Table 6, for the groups with the same contents of fat energy (36.06%) and n-6/n-3 (9 : 1), rats in EG2 fed with cholesterol level of 0.057 g/kg total diet showed significantly (P < 0.05) higher levels of TC (1.73 mmol/L) in plasma than those in EG1 fed with cholesterol level of 0.04 g/kg total diet (1.28 mmol/L). Likewise, with the same contents of fat energy (36.06%) and n-6/n-3 (5 : 1), the TC content in EG4 (cholesterol of 0.057 g/kg total diet) was 2.06 ± 0.24 mmol/L, which was significantly higher (P < 0.05) than that in EG3 (cholesterol of 0.04 g/kg total diet), 1.28 ± 0.21 mmol/L. Besides, significant difference (P < 0.05) was found between CG2 (cholesterol of 0.057 g/kg total diet) and CG1 (cholesterol of 0.04 g/kg total diet), containing 1.58 and 1.23 mmol/L of TC, respectively. Moreover, rats in EG3 taking low n-6/n-3 ratio (5 : 1) and cholesterol (0.04 g/kg total diet) showed significantly decreased TC compared to those in EG2 taking high n-6/n-3 ratio (9 : 1) and cholesterol (0.057 g/kg total diet). From these results, it is clear that no matter the fat content in the diet, the higher cholesterol feeding might cause the higher plasma TC in rats. However, TC content in EG3 fed with 5 : 1 n-6/n-3 was similar to that in EG1 fed with 9 : 1 n-6/n-3, suggesting that n-6/n-3 ratio of 5 : 1 could not reduce plasma TC levels.

Table 6

The biochemical indices of TC, TG, HDL-C, and LDL-C (mmol/L) and the ratios of HDL-C/LDL-C and HDL-C/TC.

		EG 1	EG 2	EG 3	EG 4	CG 1	CG 2
TC	B	1.16 ± 0.11	1.10 ± 0.13(a)	1.16 ± 0.20	1.16 ± 0.15(a)	1.12 ± 0.09	1.12 ± 0.12(a)
	A	1.28 ± 0.30a	1.73 ± 0.33b(b)	1.28 ± 0.21a	2.06 ± 0.24c(b)	1.23 ± 0.21a	1.58 ± 0.30b(b)
TG	B	0.48 ± 0.09	0.41 ± 0.01(b)	0.44 ± 0.03	0.53 ± 0.03(b)	0.51 ± 0.04	0.48 ± 0.08
	A	0.43 ± 0.06bc	0.35 ± 0.05a(a)	0.46 ± 0.08c	0.38 ± 0.04ab(a)	0.45 ± 0.12bc	0.44 ± 0.08bc
HDL-C	B	0.70 ± 0.07	0.69 ± 0.07	0.73 ± 0.07(b)	0.78 ± 1.04(a)	0.70 ± 0.06(b)	0.67 ± 0.10(b)
	A	0.61 ± 0.09b	0.61 ± 0.12b	0.63 ± 0.10b(a)	0.99 ± 0.11c(b)	0.54 ± 0.08ab(a)	0.50 ± 0.04a(a)
LDL-C	B	0.30 ± 0.04(a)	0.25 ± 0.07(a)	0.28 ± 0.07(a)	0.30 ± 0.09(a)	0.30 ± 0.07	0.28 ± 0.09
	A	0.39 ± 0.07a(b)	0.42 ± 0.07a(b)	0.38 ± 0.06a(b)	0.67 ± 0.11b(b)	0.36 ± 0.05a	0.39 ± 0.08a
HDL-C/LDL-C	B	2.34 ± 0.23(b)	3.12 ± 0.24(b)	2.00 ± 0.53(b)	2.73 ± 0.65(b)	2.04 ± 0.48	2.69 ± 0.32(b)
	A	1.52 ± 0.26a(a)	1.47 ± 0.29a(a)	2.24 ± 0.26b(a)	1.53 ± 0.26a(a)	2.11 ± 0.49b	1.97 ± 0.46b(a)
HDL-C/TC	B	0.61 ± 0.03(b)	0.65 ± 0.05(b)	0.65 ± 0.08(b)	0.68 ± 0.08(b)	0.67 ± 0.04(b)	0.67 ± 0.02(b)
	A	0.48 ± 0.08b(a)	0.36 ± 0.04a(a)	0.46 ± 0.07b(a)	0.45 ± 0.08b(a)	0.40 ± 0.11ab(a)	0.37 ± 0.09a(a)

Values presented as means (n = 12) ± SD. B and A represented before and after treatment, respectively. TC, total cholesterol; TG, triacylglycerols; VLDL-C, very low density lipoprotein cholesterol; HDL-C, high density lipoprotein cholesterol; LDL-C, low density lipoprotein cholesterol. Mean values within each row followed by different letters (a, b, c) are significantly (P < 0.05) different. (a)versus B and (b)versus A. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

The TG contents in EG1, EG2, EG3, and EG4 were 0.43, 0.35, 0.46, and 0.38 mmol/L, respectively. The data showed that, with the same fat energy (36.06%) for EG2/EG4 and EG3, TG contents in EG2/EG4 fed with cholesterol of 0.057 g/kg total diet were significantly lower (P < 0.05) than that in EG3 fed with cholesterol of 0.04 g/kg total diet. However, TG contents in CG1 (cholesterol of 0.04 g/kg total diet) and CG2 (cholesterol of 0.057 g/kg total diet), with the same fat energy of 30%, were 0.45 and 0.44 mmol/L, respectively, showing no significant difference (P > 0.05). There is a possibility that increased intake of cholesterol under high-fat diet could reduce plasma TG.

The contents of LDL-C in EG1, EG2, EG3, EG4, CG1, and CG2 were 0.39, 0.42, 0.28, 0.67, 0.36, and 0.39 mmol/L, respectively. The level of LDL-C in EG4 (cholesterol of 0.057 g/kg total diet) was significantly higher (P < 0.05) compared to that in EG3/EG1 (cholesterol of 0.04 g/kg total diet). It can be inferred that the increasing cholesterol content in diet could increase level of LDL-C in plasma.

With the same n-6/n-3 (9 : 1) in EG1/EG2, the content of HDL-C in both was 0.61 mmol/L, showing no significant differences (P > 0.05). Similarly, the contents of HDL-C in CG1 and CG2 (9 : 1 n-6/n-3) were 0.54 and 0.50 mmol/L, respectively, showing no significant differences (P > 0.05). However, it was 0.63 and 0.99 mmol/L in EG3 and EG4 (5 : 1 n-6/n-3), respectively, which were higher than those in EG1, EG2, CG1, and CG2 (9 : 1 n-6/n-3). Results indicated that the HDL-C level increased with the increase of n-3 PUFAs in the diet.

Furthermore, HDL-C/TC were decreased significantly (P < 0.05) in all groups after treatments. Besides, except for EG3, the ratios of HDL-C/LDL-C were also reduced significantly (P < 0.05) in all groups after treatments. With n-6/n-3 ratio of 9 : 1, fat energy of 36.06% in EG1 and EG2, or 30% in CG1 and CG2, there was no significant difference in HDL-C/LDL-C. However, with the same fat energy of 36.06% and n-6/n-3 ratio of 5 : 1, the rats in EG3 fed with cholesterol of 0.04 g/kg total diet showed significantly higher ratio of HDL-C/LDL-C (2.24 ± 0.26) than EG4 fed with cholesterol of 0.057 g/kg total diet (1.53 ± 0.26). Besides, under high-fat diet (36.06%), the ratios in EG4 (1.53 ± 0.26) were relatively higher than in EG2 (1.47 ± 0.29), but significantly higher in EG3 (2.24 ± 0.26) than in EG1 (1.52 ± 0.26), respectively, at the same level of cholesterol, difference in n-6/n-3 with 9 or 5. Moreover, rats in EG3 taking low n-6/n-3 ratio (5 : 1) and cholesterol (0.04 g/kg total diet) showed significantly increased HDL-C/LDL-C and HDL-C/TC compared to those in EG2 taking high n-6/n-3 ratio (9 : 1) and cholesterol (0.057 g/kg total diet). These results demonstrated that decreased ratio of n-6/n-3 in diet could increase the ratio of HDL-C/LDL-C in plasma of rats under high-fat diet, and the increase level was significant accompanying low cholesterol feeding.

As shown in Table 7, negative correlations were found between HDL-C/LDL-C and TC (−0.423, 0.000), between HDL-C/LDL-C and LDL-C (−0.634, 0.000), between HDL-C/LDL-C and TG (−0.539, 0.010), and between TC and LDL-C (−0.253, 0.013). Besides, a negative correlation (−0.679, 0.031) was detected between HDL-C and BWs only in EG3 (not shown in the table). However, there was a positive correlation between TC and TG (0.210, 0.042).

Table 7

The correlation analyses of biochemical indices (HDL-C/LDL-C, TC, TG, and LDL-C).

Dependent variable	Independent variable	Correlation coefficient	P value
HDL-C/LDL-C	TC	−0.423∗∗	0.000
	TG	−0.539∗	0.010
	LDL-C	−0.634∗∗	0.000
TC	TG	0.210∗	0.042
	LDL-C	−0.253∗	0.013

P lower than 0.01 (P < 0.01) was very significantly different, and P lower than 0.05 (P < 0.05) was significantly different; TC, total cholesterol; TG, triacylglycerols; VLDL-C, very low density lipoprotein cholesterol; HDL-C, high density lipoprotein cholesterol; LDL-C, low density lipoprotein cholesterol.

3.3. Effects of Dietary Fat and Cholesterol on FA Compositions in Plasma

The FAs profile in the diet could affect the FA compositions in rats' plasma (Table 8). When the n-6/n-3 PUFA ratio in the diet decreased (EG 3 and 4), the proportion of n-3 PUFA in plasma increased, thus resulting in the decrease of n-6/n-3 PUFA ratio. Besides, rats fed with n-6/n-3 ratio of 9 : 1 (EG 1 and 2; CG 1 and 2) had significantly higher level of arachidonic acid (ARA) in the plasma compared to that in rats fed with n-6/n-3 ratio of 5 : 1 (EG 3 and 4). Moreover, the proportions of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in rats feeding n-6/n-3 ratio of 5 : 1 (EG 3 and 4) were higher than those in rats feeding n-6/n-3 ratio of 9 : 1 (EG 1 and 2; CG 1 and 2). Cholesterol in diet can also affect the FA compositions in plasma. Rats fed with higher cholesterol (0.057 g/kg total diet) increased the TMUFAs (mainly oleic acid), but decreased the TSFAs (mainly stearic acid) compared to those in rats feeding cholesterol of 0.04 g/kg total diet. Further, rats in EG3 taking low n-6/n-3 ratio (5 : 1) and cholesterol (0.04 g/kg total diet) showed higher EPA and DHA and lower ARA compared to those in EG2 taking high n-6/n-3 ratio (9 : 1) and cholesterol (0.057 g/kg total diet).

Table 8

Main fatty acid compositions (% area) in rats' plasma after 60 days' treatment.

Fatty acids	EG1	EG2	EG3	EG4	CG1	CG2
14:0	0.21 ± 0.06ab	0.20 ± 0.07ab	0.16 ± 0.06a	0.21 ± 0.04b	0.21 ± 0.04ab	0.23 ± 0.06b
16:0	21.32 ± 2.08ab	22.06 ± 1.07ac	21.61 ± 1.10ab	20.72 ± 2.7a	23.21 ± 1.0c	22.68 ± 1.0bc
9c16:1	0.21 ± 0.05a	0.16 ± 0.08a	0.15 ± 0.03a	0.18 ± 0.05a	0.16 ± 0.04a	0.46 ± 0.40b
18:0	18.62 ± 2.3c	17.40 ± 1.31c	19.20 ± 3.14c	13.38 ± 4.24a	16.29 ± 2.50bc	14.63 ± 1.94ab
9t18:1	0.10 ± 0.02b	0.11 ± 0.06b	0.10 ± 0.01b	0.10 ± 0.02b	0.08 ± 0.01a	0.09 ± 0.01ab
9c18:1 OA	9.30 ± 3.06a	10.77 ± 1.61a	11.14 ± 3.05a	13.38 ± 1.94c	11.46 ± 2.29ab	13.53 ± 2.41bc
11c18:1	1.21 ± 0.22a	1.53 ± 0.31b	1.12 ± 0.24a	1.65 ± 0.27b	1.76 ± 0.42bc	1.97 ± 0.32c
9c12c18:2 LA	19.47 ± 3.76a	20.59 ± 7.03a	25.10 ± 1.56b	25.23 ± 3.22b	24.53 ± 1.92b	25.13 ± 2.15b
20:0	0.21 ± 0.06c	0.19 ± 0.02bc	0.17 ± 0.02b	0.16 ± 0.02b	0.13 ± 0.02a	0.13 ± 0.04a
18:3n-6	0.13 ± 0.06ab	0.11 ± 0.01a	0.17 ± 0.07bc	0.17 ± 0.09bc	0.15 ± 0.04abc	0.19 ± 0.06c
5c20:1	0.25 ± 0.04b	0.25 ± 0.04b	0.17 ± 0.05a	0.20 ± 0.05a	0.20 ± 0.07a	0.18 ± 0.02a
11c20:1	0.27 ± 0.12a	0.36 ± 0.10a	0.65 ± 0.25a	1.66 ± 2.09b	0.44 ± 0.18a	0.61 ± 0.20a
18:3n-3 ALA	0.39 ± 0.08ab	0.39 ± 0.08ab	0.43 ± 0.05ab	0.47 ± 0.10b	0.35 ± 0.06a	0.38 ± 0.04ab
20:2n-6	0.35 ± 0.07c	0.35 ± 0.05bc	0.23 ± 0.06a	0.31 ± 0.07bc	0.36 ± 0.10c	0.29 ± 0.03b
22:0	0.11 ± 0.06b	0.06 ± 0.01a	0.07 ± 0.03ab	0.16 ± 0.05c	0.18 ± 0.04c	0.16 ± 0.03c
20:4n-6 ARA	9.67 ± 1.90bc	11.08 ± 1.89c	7.45 ± 1.72a	7.87 ± 1.83a	10.03 ± 1.82c	9.65 ± 1.75bc
22:2n-6	0.37 ± 0.08ab	0.37 ± 0.23ab	0.26 ± 0.07a	0.37 ± 0.13b	0.33 ± 0.10ab	0.31 ± 0.09ab
20:5n-3 EPA	0.28 ± 0.02a	0.39 ± 0.06b	0.52 ± 0.05cd	0.62 ± 0.09d	0.42 ± 0.05bc	0.49 ± 0.06bc
24:0	0.11 ± 0.05b	0.06 ± 0.03a	0.05 ± 0.02a	0.06 ± 0.02a	0.05 ± 0.01a	0.04 ± 0.01a
22:3n-3	0.52 ± 0.23b	0.56 ± 0.13b	0.57 ± 0.36b	0.56 ± 0.28b	0.38 ± 0.26ab	0.18 ± 0.04a
22:4n-6	0.27 ± 0.04d	0.25 ± 0.05cd	0.15 ± 0.05a	0.19 ± 0.05ab	0.19 ± 0.05ab	0.22 ± 0.05bc
22:5n-6	0.68 ± 0.15bc	0.69 ± 0.20bc	0.74 ± 0.24bc	0.82 ± 0.19c	0.66 ± 0.15b	0.34 ± 0.11a
22:5n-3 DPA	0.50 ± 0.23	0.51 ± 0.16	0.67 ± 0.27	0.54 ± 0.12	0.45 ± 0.26	0.58 ± 0.18
22:6n-3 DHA	3.29 ± 0.42ac	3.36 ± 0.33ac	3.72 ± 0.69c	3.51 ± 0.71bc	3.02 ± 0.18a	3.07 ± 0.22ab
TSFAs	41.33 ± 3.21bc	40.82 ± 1.30bc	41.87 ± 2.67c	35.55 ± 2.23a	40.95 ± 2.47bc	38.55 ± 2.90ab
TMUFAs	11.48 ± 3.45a	13.41 ± 1.78ab	13.50 ± 3.34ab	17.78 ± 2.89d	14.25 ± 2.81bc	17.01 ± 2.84cd
TPUFAs	41.50 ± 1.09	40.20 ± 2.35	42.30 ± 1.40	42.49 ± 1.45	42.61 ± 1.15	42.20 ± 1.08
n-6 PUFAs	36.51 ± 1.14a	36.97 ± 1.21ab	36.49 ± 1.26a	36.79 ± 1.01a	37.95 ± 1.18c	37.37 ± 1.29bc
n-3 PUFAs	4.99 ± 0.69a	5.23 ± 0.54ab	5.80 ± 0.99b	5.70 ± 0.99b	4.66 ± 0.72a	4.83 ± 0.64a
n-6/n-3 ratio	7.32 ± 0.12b	7.07 ± 0.18ab	6.29 ± 0.13a	6.45 ± 0.10a	8.14 ± 0.19c	7.72 ± 0.15bc
S/M/P	1.09 : 0.30 : 1	1.01 : 0.33 : 1	1 : 0.32 : 1.01	1 : 0.50 : 1.19	1 : 0.35 : 1.04	1 : 0.44 : 1.09

Values presented as means (n = 12) ± SD. Mean values within each row followed by different letters (a, b, c, etc.) are significantly (P < 0.05) different. OA, oleic acid; LA, linoleic acid; ALA, alpha-linolenic acid; EPA, eicosapntemacnioc acid; ARA, arachidonic acid; DPA, docosapentaenoic acid; DHA, docosahexenoic acid; TSFAs, total saturated fatty acids, the sum of (C14:0 + C16:0 + C18:0 + C20:0 + C22:0 +C24:0 + ⋯); TMUFAs, total monounsaturated fatty acids, the sum of (C16:1 + C18:1 + C20:1 + ⋯); TPUFAs, total polyunsaturated fatty acids, the sum of (C18:2n-6 + C18:3n-6 + C18:3n-3 + C20:2n-6 + 20:4n-6 + C22:2n-6 + C20:5n-3 + C22:3n-3 + C22:4n-6 + C22:5n-6 + C22:5n-3 + C22:6n-3 + ⋯); S/M/P: TSFA/TMUFA/TPUFA. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

3.4. Effects of Dietary Fat and Cholesterol on FA Compositions in Liver and Muscle

The FA compositions in rats' liver and muscle are shown in Table 9. In liver, rats fed with lower n-6/n-3 ratio of 5 : 1 (EG 3 and 4) had significantly higher level of n-3 PUFA (mainly of ALA) and lower ratio of n-6/n-3 compared to rats fed with n-6/n-3 ratio of 9 : 1 (EG 1 and 2; CG 1 and 2). Similarly, the changes of n-3 PUFA and n-6/n-3 ratio in muscle showed the same tendency with liver. Interestingly, the concentrations of α-linolenic acid (ALA) in liver and muscle were higher than those in plasma, brain, and heart tissues. However, traces of EPA and DHA were registered in muscle compared to those in other organs.

Table 9

Fatty acid compositions (% area) in rats' liver and muscle after 60 days' treatment.

Fatty acids	EG1	EG2	EG3	EG4	CG1	CG2
Fatty acid compositions (% area) in rats' liver
16:0	17.41 ± 1.08a	18.19 ± 1.54ab	17.72 ± 0.82a	17.69 ± 2.90a	19.59 ± 1.86b	18.67 ± 0.90ab
9c16:1	0.43 ± 0.19a	0.58 ± 0.36bc	0.40 ± 0.12a	0.61 ± 0.37bc	0.78 ± 0.52c	0.50 ± 0.31ab
18:0	6.41 ± 1.42ab	6.49 ± 1.42ab	6.51 ± 2.08ab	7.02 ± 5.66ab	4.92 ± 2.42a	7.77 ± 3.51b
9t18:1	0.23 ± 0.02	0.23 ± 0.02	0.24 ± 0.02	0.23 ± 0.04	0.23 ± 0.02	0.22 ± 0.02
11t18:1	0.19 ± 0.02a	0.19 ± 0.02a	0.22 ± 0.02b	0.20 ± 0.04ab	0.18 ± 0.01a	0.19 ± 0.02a
9c18:1 OA	25.09 ± 2.40ab	25.02 ± 1.43ab	24.79 ± 2.40ab	25.06 ± 6.57ab	27.15 ± 3.79b	23.15 ± 4.52a
11c18:1	1.29 ± 0.18ab	1.37 ± 0.30b	1.07 ± 0.13a	1.37 ± 0.32b	1.69 ± 0.38c	1.45 ± 0.33b
9c12c18:2 LA	30.58 ± 1.33ab	29.47 ± 1.80ab	31.00 ± 1.32b	29.07 ± 2.47a	30.08 ± 2.28a	29.59 ± 2.54ab
20:0	0.11 ± 0.02b	0.10 ± 0.03ab	0.10 ± 0.03ab	0.11 ± 0.03b	0.09 ± 0.02a	0.11 ± 0.33b
18:3n-6	0.41 ± 0.10a	0.39 ± 0.06a	0.77 ± 0.23b	0.40 ± 0.07a	0.37 ± 0.07a	0.36 ± 0.08a
11c20:1	0.48 ± 0.10c	0.43 ± 0.11bc	0.28 ± 0.09a	0.35 ± 0.11ab	0.43 ± 0.13bc	0.36 ± 0.11ab
18:3n-3 ALA	1.40 ± 0.18a	1.43 ± 0.28a	3.19 ± 0.82b	2.80 ± 0.92b	1.45 ± 0.23a	1.12 ± 0.33a
20:2n-6	0.54 ± 0.12d	0.48 ± 0.10cd	0.27 ± 0.10a	0.38 ± 0.08b	0.49 ± 0.14cd	0.40 ± 0.07bc
22:0	0.04 ± 0.01	0.04 ± 0.01	0.04 ± 0.02	0.05 ± 0.02	0.04 ± 0.01	0.05 ± 0.01
20:3n-6	1.27 ± 0.22c	1.20 ± 0.20bc	1.57 ± 0.24d	1.00 ± 0.18b	0.99 ± 0.16a	1.04 ± 0.25ab
20:3n-3	0.04 ± 0.01	0.04 ± 0.00ab	0.05 ± 0.01b	0.07 ± 0.03c	0.05 ± 0.01b	0.03 ± 0.01a
20:4n-6 ARA	5.31 ± 0.55bc	5.70 ± 0.70c	3.93 ± 0.80a	4.31 ± 0.53ab	5.68 ± 0.71c	6.15 ± 0.73c
20:5n-3 EPA	0.54 ± 0.06a	0.53 ± 0.05a	0.74 ± 0.07c	0.70 ± 0.05c	0.53 ± 0.08a	0.61 ± 0.05ab
22:3n-3	0.22 ± 0.06ab	0.28 ± 0.16ac	0.21 ± 0.15a	0.32 ± 0.09ac	0.34 ± 0.18bc	0.36 ± 0.15c
22:4n-6	0.46 ± 0.12d	0.36 ± 0.08c	0.21 ± 0.08a	0.24 ± 0.06ab	0.31 ± 0.08bc	0.33 ± 0.10c
22:5n-6	0.18 ± 0.05b	0.16 ± 0.04ab	0.14 ± 0.05ab	0.13 ± 0.07a	0.13 ± 0.03a	0.12 ± 0.03a
22:5n-3 DPA	1.23 ± 0.27	1.09 ± 0.22	1.07 ± 0.29	1.16 ± 0.32	1.04 ± 0.18	1.17 ± 0.37
22:6n-3 DHA	3.93 ± 0.43	3.96 ± 0.61	3.38 ± 0.57	3.68 ± 0.73	3.08 ± 0.72	3.85 ± 0.99
TSFAs	24.70 ± 1.40a	25.75 ± 1.22ab	25.12 ± 2.33a	25.56 ± 3.16ab	25.47 ± 2.71ab	27.32 ± 3.24b
TMUFAs	27.74 ± 2.62ab	27.84 ± 1.81ab	27.00 ± 2.43ab	27.86 ± 6.78ab	30.45 ± 4.42b	25.89 ± 5.02a
TPUFAs	46.25 ± 2.18b	45.07 ± 2.21ab	46.18 ± 1.17b	45.10 ± 4.52ab	42.74 ± 3.21a	45.54 ± 2.78b
n-6 PUFAs	38.91 ± 1.68b	37.75 ± 1.78ab	37.94 ± 0.96ab	36.46 ± 3.07a	36.27 ± 2.52a	38.41 ± 1.99b
n-3 PUFAs	7.35 ± 0.61bc	7.32 ± 0.60bc	8.63 ± 0.57d	8.63 ± 1.59d	6.48 ± 0.82a	7.13 ± 1.04ab
n-6/n-3 ratio	5.32 ± 0.34bc	5.17 ± 0.30b	4.40 ± 0.33a	4.28 ± 0.43a	5.64 ± 0.46c	5.50 ± 0.86bc
S/M/P	1 : 1.12 : 1.67	1 : 1.08 : 1.75	1 : 1.07 : 1.84	1 : 1.09 : 1.76	1 : 1.20 : 1.68	1 : 0.95 : 1.67
Fatty acid compositions (% area) in rats' muscle
16:0	23.30 ± 1.90bd	22.45 ± 1.62ab	22.06 ± 0.90a	22.64 ± 1.2ac	24.00 ± 1.05d	23.74 ± 1.22cd
18:0	4.05 ± 2.42	3.96 ± 1.28	4.04 ± 0.69	3.52 ± 0.86	4.00 ± 1.36	4.07 ± 0.64
9c18:1 OA	32.57 ± 7.02	33.96 ± 4.12	35.12 ± 2.12	35.90 ± 2.33	34.28 ± 3.84	34.04 ± 2.35
11c18:1	2.10 ± 0.34c	2.17 ± 0.19c	1.57 ± 0.15a	1.75 ± 0.11ab	1.68 ± 0.78a	2.02 ± 0.11bc
9c12c18:2 LA	24.81 ± 2.04cd	25.28 ± 2.45d	22.64 ± 1.77ab	21.42 ± 1.51a	22.08 ± 2.04a	23.16 ± 1.92bc
18:3n-6	0.03 ± 0.01	0.03 ± 0.01	0.04 ± 0.01	0.03 ± 0.01	0.03 ± 0.01	0.03 ± 0.01
18:3n-3 ALA	1.42 ± 0.29a	1.40 ± 0.22a	2.93 ± 0.56c	2.53 ± 0.4b	1.27 ± 0.24a	1.28 ± 0.20a
20:2n-6	0.15 ± 0.26	0.08 ± 0.01	0.07 ± 0.01	0.07 ± 0.01	0.07 ± 0.02	0.07 ± 0.01
20:3n-6	0.25 ± 0.26b	0.20 ± 0.15ab	0.22 ± 0.11ab	0.12 ± 0.08ab	0.17 ± 0.14ab	0.09 ± 0.07a
20:4n-6 ARA	1.34 ± 0.47	1.60 ± 0.46	1.16 ± 0.40	0.90 ± 0.10	1.63 ± 0.16	1.50 ± 0.41
20:5n-3 EPA	0.05 ± 0.04	0.06 ± 0.02	0.06 ± 0.02	0.05 ± 0.03	0.06 ± 0.04	0.06 ± 0.02
22:4n-6	0.07 ± 0.07	0.06 ± 0.05	0.04 ± 0.02	0.04 ± 0.04	0.05 ± 0.05	0.04 ± 0.02
22:5n-3 DPA	0.12 ± 0.07	0.19 ± 0.06	0.21 ± 0.07	0.17 ± 0.03	0.16 ± 0.07	0.18 ± 0.04
22:6n-3 DHA	0.76 ± 0.26	0.80 ± 0.20	0.88 ± 0.23	0.90 ± 0.24	0.81 ± 0.29	0.75 ± 0.27
TSFAs	28.73 ± 4.01ab	27.87 ± 2.45ab	27.61 ± 1.40a	27.67 ± 1.50a	29.62 ± 1.87b	29.43 ± 1.45ab
TMUFAs	38.76 ± 3.18a	40.02 ± 4.41ab	42.85 ± 2.03bc	44.41 ± 3.11c	42.17 ± 4.81ac	41.71 ± 3.08ac
TPUFAs	28.92 ± 2.89	29.87 ± 2.60	28.33 ± 1.78	26.14 ± 2.99	26.38 ± 3.73	27.17 ± 2.95
n-6 PUFAs	26.56 ± 2.84bc	27.35 ± 2.79c	24.21 ± 1.52ab	22.60 ± 2.40a	24.03 ± 2.06ab	24.91 ± 1.39ab
n-3 PUFAs	2.32 ± 0.23a	2.45 ± 0.22a	4.10 ± 0.22b	3.51 ± 0.23b	2.29 ± 0.16a	2.22 ± 0.17a
n-6/n-3 ratio	11.43 ± 1.62b	11.15 ± 1.58b	5.91 ± 1.26a	6.44 ± 1.66a	10.47 ± 1.70b	11.23 ± 1.48b
S/M/P	1 : 1.35 : 1.0	1 : 1.44 : 1.07	1 : 1.55 : 1.02	1 : 1.60 : 0.94	1 : 1.42 : 0.89	1 : 1.42 : 0.92

Values presented as means (n = 12) ± SD. Mean values within each row followed by different letters (a, b, c, etc.) are significantly (P < 0.05) different. OA, oleic acid; LA, linoleic acid; ALA, alpha-linolenic acid; EPA, eicosapntemacnioc acid; ARA, arachidonic acid; DPA, docosapentaenoic acid; DHA, docosahexenoic acid; TSFAs, total saturated fatty acids, the sum of (C16:0 + C18:0 + C20:0 + C22:0 + ⋯); TMUFAs, total monounsaturated fatty acids, the sum of (C16:1 + C18:1 + C20:1 + C22:1 + C24:1 + ⋯); TPUFAs, total polyunsaturated fatty acids, the sum of (C18:2n-6 + C18:3n-3 + C20:2n-6 + C20:3n-6 + C20:4n-6 + C22:2n-6 + C20:5n-3 + C22:4n-6 + C22:5n-6 + C22:5n-3 + 22:6n-3 + ⋯); S/M/P: TSFAs/TMUFAs/TPUFAs. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

3.5. Effects of Dietary Fat and Cholesterol on FA Compositions in Brain and Heart

The FAs compositions in rats' brain and heart are presented in Table 10. Brain, a unique organ, can accumulate high concentration of DHA because this fatty acid plays a vital role in development and function of nervous system. In this study, ALA in brain was significantly metabolized to DHA as evidenced by the DHA levels in six groups varied from 11.49% to 13.66%. Interestingly, n-6/n-3 ratios in six groups were approximately with value of 1 : 1. Rats in groups fed with different cholesterol of 0.057 or 0.04 g/kg total diet showed no obvious change of FA compositions. Also, rats in EG3 taking low n-6/n-3 ratio (5 : 1) and cholesterol (0.04 g/kg total diet) showed no significant differences in EPA and DHA and n-6/n-3 ratio compared to those in EG2 taking high n-6/n-3 ratio (9 : 1) and cholesterol (0.057 g/kg total diet).

Table 10

Fatty acid compositions (% area) in rats' brain and heart after 60 days' treatment.

Fatty acids	EG1	EG2	EG3	EG4	CG1	CG2
Fatty acid compositions (% area) in rats' brain
16:00	21.53 ± 1.35	21.68 ± 1.16	21.36 ± 0.76	21.48 ± 1.05	21.09 ± 1.06	20.83 ± 0.79
9c16:1	0.38 ± 0.03ab	0.34 ± 0.03a	0.37 ± 0.04ab	0.36 ± 0.04ab	0.38 ± 0.03b	0.38 ± 0.03b
18:00	20.69 ± 0.60a	21.31 ± 0.46b	20.45 ± 0.52a	20.51 ± 0.56a	20.54 ± 0.42a	20.46 ± 0.44a
9t18:1	0.18 ± 0.04b	0.13 ± 0.12a	0.14 ± 0.03a	0.14 ± 0.02a	0.16 ± 0.03ab	0.19 ± 0.04b
9c18:1 OA	20.01 ± 1.26ab	19.22 ± 1.26a	19.95 ± 1.01ab	20.64 ± 1.11b	20.85 ± 1.05bc	21.68 ± 1.12c
11c18:1	3.86 ± 0.36abc	3.78 ± 0.31ab	3.73 ± 0.28a	4.04 ± 0.38bc	4.15 ± 0.31c	4.14 ± 0.33c
9c12c18:2 LA	1.20 ± 0.24a	1.08 ± 0.11a	1.45 ± 0.27b	1.15 ± 0.23a	1.13 ± 0.08a	1.13 ± 0.16a
20:00	0.40 ± 0.10bc	0.29 ± 0.09a	0.34 ± 0.08ab	0.34 ± 0.08ab	0.39 ± 0.09bc	0.44 ± 0.10c
5c20:1	0.06 ± 0.02ab	0.05 ± 0.01a	0.04 ± 0.01a	0.05 ± 0.00ab	0.05 ± 0.01ab	0.06 ± 0.01b
11c20:1	2.18 ± 0.69ab	1.73 ± 0.62a	2.13 ± 0.58ab	2.18 ± 0.64ab	2.34 ± 0.59b	2.50 ± 0.64b
18:3n-3 ALA	0.60 ± 0.18	0.54 ± 0.16	0.57 ± 0.15	0.59 ± 0.11	0.65 ± 0.16	0.69 ± 0.15
20:2n-6	0.23 ± 0.05bc	0.19 ± 0.04a	0.26 ± 0.04c	0.23 ± 0.06bc	0.22 ± 0.04b	0.21 ± 0.03ab
22:00	0.28 ± 0.05bc	0.23 ± 0.06a	0.24 ± 0.04ab	0.24 ± 0.03ab	0.29 ± 0.06cd	0.33 ± 0.05d
20:3n-6	0.47 ± 0.02ab	0.45 ± 0.03a	0.63 ± 0.03c	0.47 ± 0.03ab	0.48 ± 0.04b	0.46 ± 0.03ab
22:1n-9	0.24 ± 0.07bc	0.18 ± 0.06a	0.22 ± 0.06ab	0.21 ± 0.04ab	0.25 ± 0.06bc	0.27 ± 0.06c
20:4n-6 ARA	9.87 ± 0.55ab	10.28 ± 0.48b	9.81 ± 0.46a	9.66 ± 0.66a	9.53 ± 0.54a	9.45 ± 0.51a
22:2n-6	0.07 ± 0.05ab	0.04 ± 0.02a	0.06 ± 0.04ab	0.06 ± 0.02ab	0.08 ± 0.03b	0.08 ± 0.03b
20:5n-3 EPA	0.23 ± 0.04ab	0.19 ± 0.03a	0.23 ± 0.04ab	0.22 ± 0.02ab	0.24 ± 0.05bc	0.27 ± 0.06c
24:1n-9	0.12 ± 0.04ab	0.10 ± 0.03a	0.11 ± 0.02	0.11 ± 0.01ab	0.13 ± 0.03bc	0.15 ± 0.05c
22:4n-6	2.86 ± 0.19	2.81 ± 0.20	2.73 ± 0.19	2.76 ± 0.22	2.75 ± 0.19	2.77 ± 0.16
22:5n-6	0.48 ± 0.08c	0.47 ± 0.12c	0.31 ± 0.04a	0.38 ± 0.07b	0.39 ± 0.06b	0.43 ± 0.07bc
22:5n-3 DPA	0.17 ± 0.02ab	0.15 ± 0.01a	0.22 ± 0.02c	0.18 ± 0.02b	0.16 ± 0.02a	0.17 ± 0.03ab
22:6n-3 DHA	12.69 ± 1.00bc	13.66 ± 0.96d	13.26 ± 0.99cd	12.66 ± 0.73bc	12.34 ± 0.86b	11.49 ± 0.95a
TSFAs	43.33 ± 1.34ab	43.91 ± 1.33b	42.87 ± 0.93a	43.00 ± 1.10ab	42.76 ± 1.18a	42.53 ± 1.06a
TMUFAs	27.08 ± 2.33ab	25.61 ± 2.21a	26.79 ± 1.88ab	27.82 ± 2.12bc	28.41 ± 1.99bc	29.47 ± 2.02c
TPUFAs	28.99 ± 1.24bc	29.94 ± 1.09c	29.71 ± 1.12c	28.56 ± 1.13b	28.18 ± 1.16ab	27.37 ± 1.17a
n-6 PUFAs	15.16 ± 0.57bc	15.33 ± 0.48c	15.27 ± 0.48c	14.74 ± 0.67ab	14.60 ± 0.66a	14.54 ± 0.58a
n-3 PUFAs	13.83 ± 0.76bc	14.67 ± 0.78d	14.44 ± 0.79cd	13.82 ± 0.5b	13.58 ± 0.62b	12.83 ± 0.74a
n-6/n-3 ratio	1.10 ± 0.04bc	1.04 ± 0.05a	1.06 ± 0.05ab	1.06 ± 0.03ab	1.07 ± 0.04ab	1.13 ± 0.05c
S/M/P	1.49 : 0.93 : 1	1.47 : 0.86 : 1	1.44 : 0.90 : 1	1.51 : 0.97 : 1	1.52 : 1.00 : 1	1.55 : 1.08 : 1
Fatty acid compositions (% area) in rats' heart
16:00	12.68 ± 1.03bc	12.18 ± 0.33ab	13.13 ± 0.71c	11.92 ± 0.88a	12.50 ± 0.54b	12.11 ± 0.53ab
9c16:1	0.29 ± 0.54	0.11 ± 0.05	0.13 ± 0.05	0.14 ± 0.08	0.15 ± 0.07	0.11 ± 0.04
18:00	22.41 ± 1.62b	23.28 ± 0.56c	20.90 ± 0.85a	23.29 ± 0.76c	22.47 ± 0.62b	22.93 ± 0.94bc
9t18:1	0.15 ± 0.02b	0.14 ± 0.01b	0.18 ± 0.02c	0.14 ± 0.02b	0.12 ± 0.01a	0.12 ± 0.01a
9c18:1 OA	6.22 ± 2.34bc	5.02 ± 0.75a	6.41 ± 1.30c	5.65 ± 1.34ac	5.27 ± 0.93ab	5.18 ± 0.86ab
11c18:1	2.21 ± 0.14b	2.18 ± 0.06b	1.99 ± 0.13a	2.27 ± 0.21b	2.52 ± 0.12	2.49 ± 0.14c
9c12c18:2 LA	20.37 ± 1.40a	20.82 ± 0.95a	25.48 ± 2.39b	21.27 ± 1.90a	21.47 ± 1.30a	20.96 ± 0.92a
20:00	0.17 ± 0.02bc	0.18 ± 0.02cd	0.20 ± 0.04d	0.20 ± 0.04d	0.16 ± 0.01a	0.14 ± 0.01a
5c20:1	0.09 ± 0.03ab	0.09 ± 0.01ab	0.10 ± 0.01b	0.09 ± 0.01ab	0.08 ± 0.01a	0.09 ± 0.01a
11c20:1	0.16 ± 0.06a	0.14 ± 0.03a	0.40 ± 0.10b	0.27 ± 0.07b	0.14 ± 0.03a	0.14 ± 0.06a
18:3n-3 ALA	0.18 ± 0.07	0.16 ± 0.05	0.14 ± 0.04	0.14 ± 0.02	0.16 ± 0.02	0.18 ± 0.06a
20:2n-6	0.20 ± 0.02b	0.19 ± 0.02ab	0.19 ± 0.03ab	0.19 ± 0.02ab	0.19 ± 0.02ab	0.18 ± 0.01a
22:00	0.13 ± 0.02cd	0.12 ± 0.02bc	0.15 ± 0.03d	0.14 ± 0.04cd	0.10 ± 0.02ab	0.09 ± 0.01a
20:3n-3	0.61 ± 0.10a	0.60 ± 0.07a	1.13 ± 0.18b	0.68 ± 0.10a	0.62 ± 0.09a	0.54 ± 0.06a
20:4n-6 ARA	18.01 ± 0.66cd	18.08 ± 0.92d	15.33 ± 0.98a	16.91 ± 0.48ab	17.27 ± 0.86bc	17.75 ± 0.75bd
20:5n-3 EPA	0.19 ± 0.03a	0.21 ± 0.03a	0.28 ± 0.05b	0.20 ± 0.03a	0.20 ± 0.02a	0.21 ± 0.03
22:4n-6	0.44 ± 0.07b	0.37 ± 0.05a	0.36 ± 0.06a	0.34 ± 0.05a	0.35 ± 0.03a	0.37 ± 0.05a
22:5n-6	0.34 ± 0.08b	0.32 ± 0.03ab	0.32 ± 0.03ab	0.29 ± 0.04a	0.30 ± 0.04a	0.29 ± 0.03a
22:5n-3 DPA	0.69 ± 0.60a	1.83 ± 0.27ab	2.03 ± 0.35b	2.04 ± 0.27b	2.03 ± 0.23b	2.03 ± 0.32b
22:6n-3 DHA	12.47 ± 1.05ab	13.03 ± 0.83ab	12.31 ± 1.06ab	12.18 ± 1.25a	13.25 ± 1.15b	13.11 ± 1.00b
TSFAs	35.80 ± 1.08b	36.12 ± 0.53b	34.74 ± 1.24a	35.88 ± 0.90b	35.62 ± 0.63b	35.66 ± 0.85b
TMUFAs	9.25 ± 2.81b	7.81 ± 0.81a	9.37 ± 1.35b	8.72 ± 1.47ab	8.40 ± 1.10ab	8.27 ± 0.91ab
TPUFAs	54.35 ± 2.09a	55.61 ± 0.53b	55.50 ± 1.31b	55.01 ± 1.42ab	55.53 ± 1.01b	55.65 ± 0.68b
n-6 PUFAs	39.76 ± 1.87a	40.39 ± 0.67a	42.80 ± 2.19b	40.45 ± 1.63a	39.88 ± 1.49a	40.10 ± 1.02a
n-3 PUFAs	14.52 ± 1.31b	15.23 ± 0.83bc	14.76 ± 1.59a	14.56 ± 1.27b	15.6 ± 1.28c	15.53 ± 0.84bc
n-6/n-3 ratio	2.75 ± 0.31a	2.66 ± 0.19a	2.89 ± 0.49b	2.80 ± 0.32a	2.57 ± 0.32a	2.59 ± 0.19a
S/M/P	1 : 0.26 : 1.52	1 : 0.22 : 1.54	1 : 0.27 : 1.60	1 : 0.24 : 1.53	1 : 0.24 : 1.56	1 : 0.23 : 0.56

Values presented as means (n = 12) ± SD. Mean values within each row followed by different letters (a, b, c, etc.) are significantly (P < 0.05) different. OA, oleic acid; LA, linoleic acid; ALA, alpha-linolenic acid; EPA, eicosapntemacnioc acid; ARA, arachidonic acid; DPA, docosapentaenoic acid; DHA, docosahexenoic acid; TSFAs, total saturated fatty acids, the sum of (C16:0 + C18:0 + C20:0 + C22:0 + ⋯); TMUFAs, total monounsaturated fatty acids, the sum of (C16:1 + C18:1 + C20:1 + ⋯); TPUFAs, total polyunsaturated fatty acids, the sum of (C18:2n-6 + C18:3n-3 + C20:2n-6 + C20:3n-6 + C20:4n-6 + C20:5n-3 + C22:4n-6 + C22:5n-6 + C22:5n-3 + 22:6n-3 + ⋯); S/M/P: TSFAs/TMUFAs/TPUFAs. Different dietary treatments for groups: EG1 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); EG2 (0.8 mL blended oil, 9% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1); EG3 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 5 : 1); EG4 (0.8 mL blended oil, 8.6% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 5 : 1); CG1 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.04 g/kg total diet, and n-6/n-3 9 : 1); CG2 (0.6 mL blended oil, 6.1% palm oil, cholesterol of 0.057 g/kg total diet, and n-6/n-3 9 : 1). Different proportions of blended oil (g/100 g) for groups: EG1/EG2 (camellia oil 10 g, sunflower oil 74 g, and linseed oil 16 g); EG3/EG4 (camellia oil 18.6 g, sunflower oil 53.5 g, and linseed oil 27.9 g); CG1/CG2 (camellia oil 10.2 g, sunflower oil 73.5 g, and linseed oil 16.3 g).

Heart also showed high DHA levels in six groups changing from 12.18% to 13.25%. Besides, the ARA level in heart changed from 15.33% to 18.08%, higher than those in plasma, liver, brain, and muscle. However, traces of ALA were registered in heart. Similar to the results in plasma, rats fed with n-6/n-3 ratio of 9 : 1 (EG 1 and 2; CG 1 and 2) had higher level of ARA in the heart compared to that in rats fed with n-6/n-3 ratio of 5 : 1 (EG 3 and 4). And the higher cholesterol (0.057 g/kg total diet) in diet also increased the total monounsaturated fatty acid (mainly oleic acid) but decreased the total saturated fatty acids (mainly stearic acid) in heart compared to those in rats feeding cholesterol of 0.04 g/kg total diet.

4. Discussion

Dietary modification has led to an imbalance in the n-6/n-3 PUFA ratio with an increase in n-6 FAs but a marked reduction in n-3 FAs in diets, resulting in a high n-6/n-3 PUFA ratio [17]. The n-6/n-3 PUFA ratio of 9 : 1 in today's typical Chinese diets is quite different from the n-6/n-3 PUFA ratio of 1 : 1, which was found in human diets many years ago [18]. Therefore, there is necessity to investigate the effects of Chinese dietary pattern of fat content, n-6/n-3 ratio, and cholesterol content on lipid profile and FA compositions in plasma and tissues using a rat model.

Results showed that high-fat diet contributed to the increasing BWs of rats. However, rats fed with high fat, modulating by cholesterol level of 0.04 g/kg total diet and n-6/n-3 ratio of 5 : 1, showed significant decrease in BWs compared to those of rat fed with cholesterol of 0.057 g/kg total diet and n-6/n-3 ratio of 9 : 1, which indicated that, even under high-fat diet, the increase of rat's BWs could be controlled by simultaneously keeping cholesterol and n-6/n-3 at low levels. Supporting our results, Buckley and Howe [19] reported that a dietary intake of n-3 PUFAs may reduce BWs in obese rats and body fat accumulation, specifically visceral fat of rats fed with a high-fat diet. Guelzim et al. [20] showed that n-3 PUFAs could improve the body composition and insulin sensitivity and resulted in rats' weight loss. Haimeur et al. [21] also found that rats fed with high-fat diet had higher adiposity and adipose tissue/BW ratio compared to those in the control group, but these values were reduced by a dietary supplementation of n-3 PUFA.

The plasma levels of TC, TG, HDL-C, and LDL-C in rats varied with diet. The higher cholesterol fed under high-fat diet, the higher TC and LDL-C levels, and the lower TG in rats' plasma. However, under high-fat diet, the HDL-C level and the ratios of HDL-C/LDL-C and HDL-C/TC in rats' plasma increased in response to simultaneous intake of low n-6/n-3 (5 : 1) and cholesterol (0.04 g/kg total diet) compared to those of rats fed with high cholesterol of 0.057 g/kg total diet and n-6/n-3 ratio of 9 : 1. The result was in concurrence with an earlier report that cholesterol feeding in rats could significantly increase the plasma cholesterol levels, whereas it could reduce the levels of plasma TG and phospholipids [11]. Moreover, in our study, we found that, under high-fat diet, the decreased ratio of n-6/n-3 (5 : 1) in diet could not reduce plasma TC levels, which was different from the results of Jeffery et al. [22] reporting a decreased serum cholesterol concentrations as the n-6/n-3 PUFA ratio in the diet decreased. It has been reported that the incidence of atherosclerosis [23] and cardiovascular disease [24] was in close correlation with the high levels of TC, TG, and LDL-C. On the contrary, a high level of HDL-C was cardioprotective [25]. Moreover, among the possible mechanisms that may contribute to the cardiovascular benefits, n-3 PUFAs other than n-6 PUFAs were found to have the ability of decreasing the contents of TG and LDL-C with moderate increase in the level of HDL-C [26-28]. Thus, our study suggested that the decreased levels of TC, TG, and LDL-C and the increased levels of HDL-C/LDL-C and HDL-C/TC after the simultaneous intakes of low cholesterol (0.04 g/kg total diet) and n-6/n-3 (5 : 1) may be in favor of reducing the incidence of associated metabolic diseases.

Besides, we found that the accumulation of various FAs was associated with the dietary intake of n-6/n-3 ratio, cholesterol, and even tissues. Although the blended oils fed to rats were with no EPA and DHA, these two PUFAs were detected in rats' plasma and four organs, especially high in brain and heart. This is because dietary ALA can convert into long-chain n-3 PUFA, such as EPA, DPA, and DHA due to the desaturation, elongation, and β-oxidation reactions [29, 30]. Increased levels of EPA, DHA, and ALA and decreased level of ARA were observed in rats' plasma after the intake of n-6/n-3 (5 : 1) and cholesterol (0.04 g/kg total diet) compared to rats fed with n-6/n-3 ratio of 9 : 1 and cholesterol of 0.057 g/kg total diet. The result suggested that negative correlations were found between the levels of DHA, ALA, or EPA and ARA in plasma. This may be due to the antagonistic effect between DHA, ALA, or EPA and ARA in absorption and metabolism. Moreover, as the n-6/n-3 PUFA ratio of the diet decreased, the proportion of n-3 PUFA increased in plasma, liver, and muscle, thus resulting in the decreased n-6/n-3 PUFA ratio. In accord with our results, Jeffery et al. [31] found that there were decreases in the levels of linoleic acid and ARA and an increase level of ALA in tissues as the n-6/n-3 ratio of the diet decreased. Umesha and Naidu [32] reported that dietary feeding of blended oils with n-6/n-3 ratio of 2.3–2.6 significantly increased ALA, EPA, and DHA content in serum, liver, and other tissues. We also found that the increasing cholesterol in diet increased the TMUFAs but decreased the TSFAs in rats' plasma and heart, which is in agreement with the results of Huang et al. [33] reporting an increase of 18:1n-9 after feeding rats with cholesterol. Therefore, due to the beneficial effect of n-3 PUFAs against n-6 PUFAs on metabolism, it may be helpful to prevent diseases by controlling dietary intakes of n-6/n-3 ratio (improving n-3 PUFAs) and cholesterol level at a low level.

5. Conclusions

High-fat diet increased the BWs of rats; however, the increase could be controlled by intakes of low cholesterol (0.04 g/kg total diet) and n-6/n-3 ratio (5 : 1). Besides, plasma lipid levels (TC and LDL-C) were also increased and HDL-C level was decreased under high-fat or high-cholesterol diet, but they were ameliorated in response to simultaneous intakes of low n-6/n-3 (5 : 1) and cholesterol (0.04 g/kg total diet). Likewise, the proportion of n-3 PUFA increased and the ratios of n-6/n-3 decreased in plasma, liver, and muscle as the n-6/n-3 PUFA ratio in the diet decreased.