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Animal nutrition

Straight from the cow, diet milk

Molecule added to feed reduces level of fat, increases proteins, and intensifies anti-cancer effect

With infinite bovine calm, a group of cows is feeding on grass and animal feed in the stables on Embrapa’s experimental farm in Vassouras, RJ. They are special: their milk has 30% less fat and 10% more protein, as well as anti-cancer properties. In three years, Brazilian cattle breeders will probably be able to distribute this milk, which has been achieved, following studies at Esalq, the Luiz de Queiroz Agriculture College, of the University of São Paulo (USP), in Piracicaba.

This milk, equivalent to semi-skimmed, is the most recent experiment by Esalq’s Dante Pazzanese Lanna, who has worked in partnership with researchers from the United States and from Embrapa, the Brazilian Agricultural Research Corporation. This month, an engineer in agronomy, Sérgio Raposo de Medeiros, whose doctorate Lanna is supervising, is presenting the good news at a meeting of the American Society of Animal Science, in the USA. In August, it is Lanna’s turn; he will be showing it at the Congress of the European Association of Animal Production, at The Hague, in Holland.

To obtain the milk that nutritionists dream of, Lanna prepared a feed with modified molecules of CLA, conjugated linoleic acid, a fatty acid that forms lipids (fats).  Added to the animal’s food, the modified acid acts as a pharmaceutical compound, altering the synthesis of fat in the mammary gland.

It has been s four years since Lanna started studying CLA’s properties, which is produced naturally in the stomachs of ruminants (cattle, sheep, and goats), and found in food of animal origin, above all in milk. Its biological and anti-cancer effects have been reiterated in international research. In 1996, the National Council of Sciences of the United States confirmed that CLA is capable of inhibiting tumor formation. Besides preventing cancer, it attacks cancerous cells and reduces tumors that have already been formed. In experiments on mice in the Roswell Park Institute, in New York, United States, CLA brought about a significant reduction in the number of mammary tumors, even in very low concentrations. With doses in the order of 0.25% of the diet, it reduces by half the number of tumors, compared with animals that were not receiving it.

The CLA molecule has 18 carbon atoms and exists in several forms – isomers – in which these atoms are grouped differently, which alters their effects on the organism. One of the isomers is the CLA18:2 cis 9, trans 11 (c9, t11) – the abbreviations indicate that the CLA contains a double link in carbon 9 in the cis form, that is, on the same plane of the molecule, and another double link in carbon 11 in the trans form, that is, on another plane, as if the molecule were to turn on its own axis. American researchers recently confirmed in mice that CLA c9,t11 prevents the formation of cancerous cells.  The formation of plaques of fat (arteriosclerosis) on the walls of the blood vessels is apparently reduced by another isomer, CLA 18:2 t10,c12. “We are now trying to find out the quantity necessary to optimize protection against cancer, and to add this molecule to food consumed by the population in general”, says Esalq’s researcher.  It is known that the CLA 18:2 trans 10,cis 12 isomer inhibits the synthesis of fat in milk, and it therefore has an anti-obesity effect.

Lanna imagined a challenge: could it be possible to raise the level of protein in milk?  The greatest difficulty in this was to obtain a combination of conjugated linoleic acid that contains, in particular, the t10, c12 isomer, added to an amino acid supplement.  The hypothesis that more amino acids would have to be put into the cow’s diet, for it to produce more casein, (the protein in milk) proved correct, and a noteworthy increase in the level of protein resulted. Lanna’s team patented the effect of CLA in reducing fat in milk. Now, researchers from Embrapa, Esalq, the University of Idaho, and FAPESP are applying for the registration of the discovery that CLA increases the level and production of protein in milk by up to 10% and 20% respectively.  It is not their only unprecedented achievement. “We have also demonstrated, for the first time, an increase in the production of milk in the pasture, and an increase in the duration of lactation, which makes the cow keep up a high production of milk during the lactation period of about ten months”, he says.

Now in contact with the private sector, the researchers want to offer a low cost supplement, which appears to be viable, because linoleic acid, CLA’s raw material, can easily be obtained from foods like olive and soybean oil. It may be a way to expand both the production of milk (which is 20 billion liters today), and its consumption in Brazil, estimated at 137 liters per inhabitant / year, the equivalent of a glass a day, or one third of what is recommendable to avoid problems like osteoporosis (please see Pesquisa FAPESP 52).

Grandfather’s influence
With his work, Lanna is following the steps of his maternal grandfather, the doctor from Minas Gerais, Dante Pazzanese, who dedicated his life to the treatment and cure of heart diseases. A son of Italian immigrants, he brought the first electrocardiogram equipment to Brazil, and, in 1954, he founded the Institute of Cardiology, in São Paulo, which was one of the first institutions of its kind in Brazil; today, it bears his name. When he retired in 1974, he dedicated himself to the farm in the north of Paraná, where he planted coffee and soybeans and raised cattle. The time he spent in the company of his grandson, who would spend his holidays there, was short – the doctor died in the following year, at the age of 75 – but intense, and aroused in the boy an interest for agronomy.  Another push towards university came from his father, Amadeu Duarte Lanna, a professor of anthropology at the University of São Paulo (USP), and the author of one of the first projects in the humanities financed by FAPESP, on the Indians in the Xingu National Park, in Mato Grosso.

Recently concluded, Dante Lanna’s research paper, Conjugated Linoleic Acid: Effect of Supplementation for a Long Period on Cows during Lactation, received funding of R$ 48.800 from FAPESP, and the collaboration of researchers from the universities of Cornell and Idaho, in the United States, and of Helsinki, in Finland. Right at the beginning, a heartening result came from Cornell: four dairy cows that had received a supplement to their food of 50 grams of CLA for four days had a reduction of more than 50% in the fat level in their milk. “After three days of treatment, the animals started to produce milk with less than 1.5% of fat, and so less fat than semi-skimmed milk (with 2% of fat) that is sold in the supermarkets”, comments Lanna, who corroborated the same results here, in a group of 32 animals.

On the basis of this discovery, Lanna and his former colleague at Cornell, Mark McGuirre (now with Idaho) started experimenting with the supplementation of CLA for longer periods (two months), under commercial production conditions. “We didn’t even know if the animals would have metabolic problems for producing such low-fat milk for such a time”, says Esalq’s researcher. In the United States, McGuirre worked with purebred Friesian cows with a high potential for production (over 50 kilos of milk a day), in intensive confinement conditions. Over here, Lanna and Luiz Aroeira, from Embrapa, used crossbred Friesian cows with a lower potential for production (18 kg/day) and kept in the pasture. They would eat 4 kg/day of concentrate with CLA, and a supplement of proteins to supply amino acids.

The studies led to similar results in both countries. In the initial stage, when they were trying to find out whether CLA would be harmful to the animals, it was proved that, even for prolonged periods, it did not cause any digestive, metabolic, reproductive or sanitary problem.  Later on, the two studies attested the fact that CLA increased the production of milk and kept productivity up after the start of lactation. In the Brazilian experiment, the effect of improving the duration of lactation continued, even after suspending the supply of CLA in the twelfth week.

Both in the United States and in Brazil, the initial laboratory results were confirmed, with reductions in the level of fat of around 26%. By reducing the fat in the milk, CLA also reduces the cow’s energy requirements. This makes its organism channel more nutrients towards the replacement of the fat lost at the beginning of lactation, improving its bodily health and its rate of reproduction. The cows that were the subject of research came to produce 20 liters of milk a day, even in the pasture, which supplies less energy than that obtained with a corn diet, for example.

Extra gain
The research indicates another beneficial effect: the controlled addition of CLA to the animal feed inhibits the synthesis of fatty short chain saturated acids, including myristic and lauric acids – the most dangerous for health, as they increase the levels of cholesterol. And, at the same time, it increases the proportion of long chain acids, like palmitic acid, which has less effect on the level of cholesterol, and probably reduces the chances of heart disease.

There were differences in the experiments.  In the cows treated in the United States, the level of protein in the milk increased 4%, whilst in Brazil the increase was 11%.  The reason: the animals here received larger quantities of protein and minerals than was regarded as necessary for the crossbred Friesians in lactation.  The diet that the researcher made up included fish meal and soybean bran with heat treatment, which avoids the excessive degradation of the proteins during digestion by the bacteria in the rumen (the first compartment of the bovine stomach).  And so, there is plenty of protein, which the mammary gland can use to secrete into the milk.  Besides the milk of the cows fed on this diet, in a proportion of 5 grams of CLA per kilo of feed, having an increase of 11% in the level of protein, the increase in the production of milk made the total production of protein grow 19% (see charts).

According to Lanna, the molecule of conjugated linoleic acid that inhibits the synthesis of fat by cows is the CLA t10,c12, which acts like a metabolic modifier or sharer of nutrients: it redirects the nutrients, reducing the proportion of those used in fat synthesis, and increasing those used to synthesize protein.  The two processes take place simultaneously in the cells. As it was confirmed, CLA inhibits the synthesis of fat in milk, and allows more nutrients to be redirected for the synthesis of proteins in the mamma.

There was a risk, however: the bacteria in the rumen can transform CLA into linoleic acid, which has no metabolic effects. To avoid this, the animals received in their feed CLA protected by calcium salts.  It worked: the mixture of the molecules of CLA t10,c12 and t11,c9 and the supplement with metabolizable proteins did not become degraded in the rumen; they were absorbed by the cow and altered the metabolism of the mammary gland, as expected.

In summary
The course of the project run by the engineer in agronomy and biochemist, Dante Pazzanese Lanna can be summed up in four stages:

1.         Conjugated linoleic acid (CLA) is protected from degradation in the rumen (in the first compartment of the stomach of the cow) by means of the addition of calcium salts.

2.         The CLA with calcium is added to the feed given to the cows, together with a supplement of amino acids.

3.         The isomer CLA t10,c12, absorbed by the cow’s organism, reaches the cells of its mammary glands, where it reduces the influence of the genes of the FAZ and ACC enzymes, which are responsible for the synthesis of fat in milk.

4.         The cow starts to produce milk in greater quantity, with less fat, more protein, and also containing the CLA c9,t11 isomer, which exerts a strong anti-cancer and anti-arteriosclerosis action (combating the formation of plaques of fat that can obstruct the arteries).

The discovery impact will be great. After all, this semi-skimmed milk and with its important therapeutic effects is an entirely natural product, obtained without having to undergo any industrial process. And, since milk is the raw material of many foods, the whole chain of foodstuffs should be reached.

Among the producers and processors of milk, this research deserves to be celebrated , for reviving the interest in a product whose consumption has been limited to children and adolescents.  “Over the last decade”, says the researcher, “there has been a considerable reduction in the consumption of butter, and, simultaneously, an increase in the consumption of skimmed milk, because fat has been pointed out as one of the main villains in heart diseases”.  He adds that “the research will make it possible to control the production of the components of milk, in accordance with the needs of the market”.

Stables and laboratory
Lanna is working on another project supported by FAPESP – The Effect of the Growth Hormone on the Expression of Genes in the Enzymes that Regulate Lipogenesis and Lipolysis – with finance of R$ 171,600, and its conclusion forecast for 2001. The aim, this time, is to understand the cellular and molecular mechanisms in the action of CLA and the hormones in the cows’ mammary and adipose tissue.  These mechanisms may explain in greater depth how the production of fat in milk occurs.

The reduction in the level of fat is a result of the reduced activity of certain genes. These genes codify the enzymes FAS, fatty acid synthetase and ACC, Acetyl-CoA carboxylase, which are responsible for the synthesis of fat.  The two enzymes are the motors for transforming into fat the nutrients that arrive with the blood.  Using a sample of tissue from the mamma of a cow that has received CLA, the researchers extract molecules of RNA, ribonucleic acid – which take to the remaining cells information on the genes contained in the DNA.  Alterations in the expression of these genes are determined by means of a technique that makes it possible to know how many copies of this RNA messenger are made.

First of all, Lanna’s team, in conjunction with the Xylella Genome project group at Esalq, performed the cloning and the sequencing of some of these genes, unknown in cattle.  The objective is to get to know the mechanisms involved in regulating the secretion of milk, so as, for example, to develop transgenic animals that produce more milk – with the properties that the consumer regards as ideal – and that are more efficient in the conversion of food.

And then there is researcher Sérgio Medeiros, assessing the presence of the different forms of CLA in the food consumed by the customers at Esalq’s restaurant. He wants to know how much of the substance has to be added to enrich the food and to provide protection against cancer. And zootechnician Dimas Estrásulas de Oliveira, in another project for a doctorate, assesses whether cows eat more or less grass, when they receive a supplement of CLA. His work may be capable of defining how many kilos less of feed a cow fed with CLA needs to produce a liter of milk.

The team has the same objective: to produce milk with less fat and more efficiency.  Getting closer all the time to the ideal properties and a clear understanding of how CLA works, Dante Pazzanese Lanna is trying to reduce the occurrence of heart problems – the dream of his grandfather, who taught him to like research and agronomy.

Alternatives for fattening cattle

Another research coordinated by Dante Pazzanese Lanna in Esalq’s Laboratory of Animal Nutrition and Growth, ready almost a year ago, is the Maximum Profit Feed program for bovine nutrition.  Carried out by four of Esalq’s researchers – Lanna, Luis Gustavo Barioni, Luis Orlindo Tedeschi and Celso Boin –, it won the collaboration of specialists from Cornell and R$ 6,000 in finance from FAPESP.

The coordinator calculates that the program is already being used by around 200 animal feed factories, consultants and cattle raisers in South America, involving at least 250,000 of the 1.3 million head of cattle under confinement in the country.  Until a short time ago, there was no similar model for the Brazilian reality.

The work integrates knowledge on the nutrition of cattle within a computer program, which includes a library of feeds available in Brazil. It simulates the metabolism of nutrients, the quantity of food, the protein and fat contents of the feeds, and the gain in weight.

Adapted to cattle raising in the Brazilian tropics, and for the zebu races of cattle (85% of the nation’s herd of 160 million head, the largest in the world), the program permits individualized management for each property.  For the economic analyses, it has a non-linear system of optimization put forward by Lanna and Barioni, recently published at the International Congress of Modeling and Simulation, in New Zealand.  When, for example, corn, sorghum and the pulp of citrus fruits are available, all that has to be done is to supply the price of each product, for the program to point out the most advantageous option.  Afterwards, it goes through all the possible combinations of these foodstuffs and identifies the most economical, with an estimate of the consumption and the gain in weight.  It costs R$ 180, and is distributed by Fealq, the Luiz de Queiroz Foundation of Farm Studies.

Dante Pazzanese Lanna, aged 36, became an engineer in agronomy in 1985, at Esalq, the Luiz de Queiroz Higher School of Agriculture of the University of São Paulo (USP), where he took a master’s degree, and has been lecturing since 1996.  He became a doctor in Biochemical Nutrition at Cornell University, USA, in 1993.
Projects: Conjugated Linoleic Acid: Effect of Supplementation for a Long Period on Cows during Lactation; Effect of the Growth Hormone on the Expression Genes that Regulate Lipogenesis and Lipolysis; and Development of Models for the Use of Nutrients in Ruminants and for the Determination of Nutritional Requirements
Investments: R$ 48,872.80; R$ 171,661.60; and R$ 6,000.00