Study on Carotenoid Lycopene in Animal Nutrition

Oxidative stress seriously threatens the productivity and health of livestock and poultry, causing huge economic losses to the animal husbandry industry. In the production process of livestock and poultry, factors such as environmental changes, physiological changes, and exogenous pathogenic toxins (such as mycotoxins) can cause oxidative stress, thereby disrupting the redox balance in animals. The excessive production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) can cause irreversible damage to cell lipids, proteins, and DNA, thereby affecting the physiological functions and production performance of animals.

Antioxidant protection is defined as the process of antioxidant defense in living organisms. Antioxidants in the body are mainly divided into two categories: one is synthesized by the body itself, and the other is obtained from food. Under special circumstances such as high temperatures, weaning, and pregnancy, supplementing exogenous antioxidants (such as plant-derived polyphenols and carotenoids) can effectively relieve the oxidative stress state of livestock and poultry, reduce oxidative damage, and improve the health and production performance of livestock and poultry.

Antibiotic resistance and residues have a negative impact on animal production, human health and environmental sustainability. In 2006, the European Union banned the use of antibiotics as feed additives in animal feed, and in July 2020, China officially banned the use of antibiotics in animal feed. Therefore, there is an urgent need to develop natural, green and safe alternatives to antibiotics.

Lycopene (C40H56) is a carotenoid that is a natural pigment in plants, mainly found in fruits and vegetables such as tomatoes, carrots, watermelons, and pomegranates. Lycopene has been listed as a nutrient and food additive in many countries and is widely used in food, medicine, cosmetics, agriculture, and other fields. Lycopene contains two non-conjugated double bonds and 11 conjugated double bonds in its structure, and its chemical structure gives it the greatest antioxidant capacity. Lycopene's role as a powerful antioxidant is the basis for its health-promoting effects, including anti-inflammatory, anticancer and hypoglycemic potential, cardiovascular protection, and neurobiology and hypotensive effects. An increasing number of studies have shown that lycopene can be used as a functional feed additive for livestock and poultry, and has been reported to improve production performance, meat quality, egg quality, antioxidant properties, immune function, lipid metabolism and intestinal physiological function.

I. Lycopene structure, absorption and biological functions

1. 1.Lycopene sources and structure

Among the rich carotenoid family, there are about 60 species present in food, and only 20 species can be detected in the blood and tissues of humans and animals. Due to their conjugated bonds (at least 7), they give food its color, mainly yellow and red. Differences in structure distinguish two groups of carotenoids: carotenoids (lycopene, α-carotene, β-carotene, etc.) and xanthophylls (lutein, astaxanthin, canthaxanthin, etc.).

Lycopene was discovered in tomatoes in 1876 and was named after the scientific name Lycopersicon esculentum 27 years later. Lycopene is widely found in tomatoes and also in vegetables and fruits such as carrots, sweet potatoes, pumpkins, watermelons, apricots, papayas, grapefruit and guavas. The main source of lycopene is tomatoes (80%), which are the main source of lycopene extraction and also the cheapest ingredient. The lycopene content of tomatoes varies greatly, depending on factors such as variety, maturity, climate and geographical location of cultivation. The technology for extracting lycopene from raw materials usually includes chemical extraction, microwave and ultrasound-assisted extraction, supercritical fluid extraction and enzyme-assisted extraction. A variety of biotechnologies have been developed for the large-scale production of lycopene, among which microbial fermentation is a typical traditional biotechnology in the production of lycopene. In addition, modern biotechnology, including genetic engineering, protein engineering and metabolic engineering, has also been applied to the production of lycopene.

Contrary to most carotenoids, lycopene has a linear structure. Lycopene is a polyene chain consisting of 13 double bonds, 11 of which are conjugated in a linear arrangement, making it longer than other carotenoids. Lycopene is a red waxy pigment that occurs in nature as slender needle-like crystals. Lycopene is fat-soluble, insoluble in water, and soluble in benzene, chloroform and acetone. Lycopene occurs in nature mostly in the all-trans configuration, which is relatively stable, but at least 50% of the cis isomers are found in human blood plasma and tissues. The common forms are the 5-cis, 9-cis, 13-cis and 15-cis isomers, which indicates that the cis isomers are more readily absorbed and utilized by humans and animals. Food processing is actually a value-adding step, because after heat treatment, more lycopene becomes more bioavailable. If tomato juice is exposed to cooking temperatures, cis isomers are formed, which are considered to be more bioavailable. The double bond of lycopene can be isomerized from all-trans to mono- or polycis under the influence of light, temperature or chemical reactions. Lycopene is acyclic, has a symmetrical planar structure and is particularly susceptible to oxidative degradation as a highly conjugated polyene. Physical and chemical factors such as high temperatures, exposure to light, oxygen, extreme pH values and molecules with reactive surfaces can damage the double bond of lycopene. The undesirable degradation of lycopene not only affects the sensory quality of the end product, but also the health benefits of tomato-based foods for humans and animals.

1.2. Lycopene absorption

Lycopene is the carotenoid with the highest blood plasma concentration in humans, with the average concentration depending on nutritional habits. In terms of absorption, the most widely studied is β-carotene, not lycopene and other carotenoids. Although further research is needed, the factors affecting β-carotene absorption are likely to have a similar effect on lycopene. Similar to other carotenoids, lycopene is embedded in its food matrix and cannot be absorbed efficiently by humans and animals. Lycopene is absorbed in a similar way to lipids, mainly through passive diffusion. Lycopene in the food matrix is released by the action of gastric acid, bile acids and enzymes. Once it enters the intestine, it binds to lipids to form chylomicrons, which then enter the mesenteric lymphatic system via diffusion and osmosis and finally enter the portal vein circulation. This is the main route of absorption of lycopene from the gastrointestinal tract.

After being released from the food matrix, the lycopene ingested must be emulsified and dissolved in the chylomicrons before it can be absorbed into the intestinal mucosa. Lycopene can also be absorbed via the transport protein Transporter, B-type, 1 (TBP-1), which involves an active process. The scavenger receptor class B type 1 transporter is found mainly in the small intestine, liver, adrenal glands, ovaries, placenta, kidneys, prostate and brain, and is partly responsible for transporting carotenoids from lipoproteins to tissues and from tissues to lipoproteins. Lycopene is supplied through the diet and is digested in the gastrointestinal tract by 10% to 30%. Lycopene and its metabolites are released and transported via the low-density lipoprotein and very low-density lipoprotein, and are finally distributed to target tissues. They circulate through the body and preferentially accumulate in the testes, adrenal glands, liver, prostate and adipose tissue. This uneven distribution suggests that they have unique biological functions in these tissues, such as regulating lipid metabolism in the liver.

1.3. Biological function of lycopene

Lycopene powder is one of the strongest antioxidants among carotenoids after astaxanthin. Other antioxidants include α-tocopherol, carotene, cryptoxanthin, zeaxanthin, β-carotene and lutein. Lycopene is a powerful antioxidant, which is the basis for its health-promoting effect. The most abundant carotenoids in human plasma are β-carotene, α-carotene, β-cryptoxanthin, lutein, zeaxanthin and lycopene. These six major carotenoids account for 70% of all carotenoids in human plasma and tissues. In terms of biological function, lycopene mainly acts as a singlet oxygen and peroxyl radical scavenger. Lycopene's unique double bond structure makes it far superior to other carotenoids in scavenging both human and animal peroxyl radicals. Lycopene is twice as effective as beta-carotene and 10 times more effective than alpha-tocopherol in scavenging singlet oxygen (a type of ROS). The mechanism of action of lycopene on active substances can be explained by three possible mechanisms: the formation of adducts, electron transfer to free radicals, and the extraction of allylic hydrogen. Among them, the formation of adducts, free radicals attach to the polyene chain, that is, the highly conjugated double bonds of lycopene, to form lycopene-peroxyl radical adducts.

In addition to neutralizing ROS, lycopene also activates the expression of genes encoding NAD(P)H:quinone oxidoreductase, heme oxygenase 1, glutathione reductase and glutathione S-transferase, which help to scavenge free radicals and reduce inflammatory damage. These enzymes are considered antioxidant and detoxification enzymes, also known as phase II cell protection enzymes. The promoter region of the inducible genes encoding these enzymes contains ARE, which, upon binding to Nrf2, leads to an increase in the expression of these genes. The Nrf2/ARE signaling pathway is an important endogenous antioxidant defense mechanism. Lycopene acts primarily as an anti-stress agent, relieving oxidative stress and maintaining the health of poultry by triggering the Nrf2/ARE transcription system. Keap1 is a specific receptor for the Nrf2 gene. Lycopene blocks the binding of Nrf2/Keap1, releasing Nrf2, which is then transported to the cell nucleus and upregulates the expression of phase II cell protection enzymes in heat-stressed poultry. In addition, a growing body of research has shown that lycopene has anti-inflammatory, anticancer and anti-diabetic potential. Lycopene has also been shown to have cardiovascular protective effects, neurobiological effects, and blood pressure-lowering and anti-platelet aggregation effects.

2. Application of lycopene in livestock nutrition

2.1. Effects on production performance

Sun et al. reported that a daily supplement of 50 mg/kg lycopene during gestation and lactation can improve the reproductive performance of sows, including an increase in the rate of live piglets born, the rate of live piglets weaned, the birth weight of litters, the weight of litters weaned, and a reduction in the rate of dead piglets born. The final conclusion was that lycopene can improve the reproductive performance of sows by regulating milk composition, placental immunity and antioxidant capacity. During the fattening period, dietary lycopene supplementation does not affect animal production performance. Fachinello et al. that various lycopene additive levels (12.5, 25, 37.5 and 50 mg/kg) did not affect the growth performance of finishing pigs, and that feeding finishing pigs 12.5, 25, 37.5 and 50 mg/kg of lycopene did not affect their carcass characteristics or relative organ weights.

An et al. demonstrated in a poultry nutrition study that 20 mg/kg lycopene or 1.7% tomato paste in the diet for 28 days increased egg weight and egg production in Hy-line Brown hens. Wan et al. showed that 10, 20 or 30 mg/kg lycopene increased the average daily weight gain of broilers. However, in a study by Lee et al., no positive effects of diets containing 10 or 20 mg/kg lycopene or 17 g/kg tomato paste on the growth performance and relative organ weight of broilers were observed when tomato paste was used as the lycopene source. Lycopene has been shown to have a growth-promoting effect in animals under stress conditions (e.g. heat stress) and mycotoxin feed contamination. Under heat stress conditions, the addition of 0, 200 and 400 mg/kg lycopene for 42 days in a row was also shown to linearly improve the growth performance of Ross 308 broilers, as evidenced by an increase in cumulative feed intake, an increase in body weight and a decrease in feed conversion ratio. Sarker et al. Under conditions of feed challenge with aflatoxin B1, lycopene supplementation at 100 mg/kg increased the average daily weight gain of broilers from 1 to 12 days of age, and lycopene supplementation at 200 mg/kg and 400 mg/kg increased the average daily weight gain of broilers from 22 to 42 and 1 to 42 days of age, respectively.

2.2. Effect on meat quality and egg quality

Lycopene, as a natural feed additive, has attracted widespread attention in animal production for improving meat and egg quality. Wen et al. reported that the addition of lycopene to the diet improved the meat quality of finishing pigs, including a decrease in L* and b* values, an increase in a* values, and an increase in intramuscular fat and crude protein content in the longissimus dorsi muscle. Lycopene was also shown to promote the conversion of muscle fibre types in pigs, which is important in determining meat quality. In the study by Wen et al., the addition of lycopene resulted in an upregulation of mRNA levels of myofibrillar markers such as cytochrome c, myosin heavy chain IIa and myosin heavy chain IIx in the longissimus dorsi muscle of finishing pigs. Lipid oxidation can have a negative impact on the colour, nutritional value and flavour of meat as well as its shelf life. Wen et al. also showed that the addition of lycopene improved the antioxidant status of the longissimus dorsi muscle of finishing pigs, as evidenced by increased total superoxide dismutase and catalase activities, reduced MDA levels, and up-regulated mRNA levels of SOD1, SOD2, CAT, GPX1, GST, GR and Nrf2 (all of which are oxidative stress response factors). Similar results were reported by Correia et al., who observed improved oxidative stability in the longissimus dorsi muscle of piglets after feeding 5% tomato pomace for 5 weeks. However, An et al. found that the addition of 20 mg/kg lycopene + 3.4% tomato paste or 10 mg/kg lycopene + 1.7% tomato paste to the feed had no effect on the fatty acid composition of the abdominal meat of finishing pigs in a 28-day feeding trial.

In terms of egg quality, Shevchenko et al. showed that adding lycopene (20, 40, 60 mg/kg) to the diet of High Line W36 laying hens for 90 days improved egg quality, with increased carotenoid levels and yolk color in fresh eggs and eggs stored at 4°C and 12°C. Orhan et al. showed that feeding Lohman LSL hens with 20 mg/kg lycopene pure powder or tomato powder for 84 days increased egg weight, yolk color, yolk weight, yolk-to-egg ratio, and yolk lycopene levels, and lower the levels of egg yolk MDA and cholesterol. In the study by An et al., adding 10 and 20 mg/kg lycopene to the feed for 28 days increased the egg yolk color and lycopene levels and lowered the MDA levels in Hy-line Brown eggs.