Trans fatty acids (TFA) are unsaturated fatty acids that contain at least one double bond in the trans configuration (i.e. the two hydrogen atoms are on opposite sides of the carbon to carbon double bond), resulting in a straighter shape more similar to saturated fatty acids. Consequently, TFA are less fluid and have a higher melting point than unsaturated fatty acids with cis double bonds, which are the major monounsaturated fatty acids (MUFA) and polyunsaturated fatty acids (PUFA) in plants and animals (Bhardwaj et al., 2011).

TFA in foods mainly come from two sources, including partially hydrogenated vegetable oils (i.e. industrial TFA) and ruminant-derived foods such as dairy and beef (i.e. ruminant TFA). Industrial TFA are found in partially hydrogenated vegetable oils generated using hydrogen in the presence of a catalyst (Bhardwaj et al., 2011), while ruminant TFA are made by rumen bacteria using a process called biohydrogenation (Lichtenstein, 2000; Dhaka et al., 2011).

Until recently, industrial TFA were extensively used by the food industry as a replacement for saturated fats. However, in recent years, many countries have banned industrial TFA from the food supply due to their detrimental effects on cardiovascular health (Lichtenstein, 2000; Mozaffarian et al., 2006). Recent government bans on industrial TFA in developed countries including the U.S, Canada and the E.U has left ruminant-derived fats (e.g. dairy, beef, lamb and goat) as the sole source of TFA in the food supply. As such, there is heightened interest in the composition, content, and health effects of ruminant TFA particularly conjugated linoleic acid (CLA) and trans 18:1 isomers (Brouwer et al., 2010; Prada et al., 2022). The aim of this review is to provide an update of the current state of knowledge on the biosynthesis, concentration range and factors affecting the concentration of TFA in ruminant fats, current intake of ruminant TFA, and to evaluate evidence on their health effects with an emphasis on type 2 diabetes.

Ruminant TFA are formed via biohydrogenation of unsaturated fatty acids in the rumen. The process of biohydrogenation is performed by rumen bacteria, during which dietary unsaturated fatty acids are converted to saturated fatty acids (Lichtenstein, 2000; Dhaka et al., 2011; Dugan et al., 2018). Dietary unsaturated fatty acids are toxic to rumen microbes, hence they biohydrogenate them to saturated fatty acids which are neutral or less toxic (Jenkins et al., 2008). The biohydrogenation process includes several isomerization and hydrogenation steps which result in the formation of many intermediates including conjugated and non-conjugated trienoic, dienoic and monoenoic trans fatty acids (Jenkins et al., 2008; Vahmani et al., 2015). A small portion of these intermediates passes the rumen and subsequently find their way into tissues and milk after post-ruminal absorption (Lichtenstein, 2000; Dugan et al., 2018).

The predominant fatty acids in ruminant diets include 18:2n-6 (linoleic acid; LNA) and 18:3n-3 (alpha-linolenic acid; ALA) and thus are considered the main substrates for ruminal biohydrogenation. It is estimated that on average, about 80% and 92% of dietary LNA and ALA are biohydrogenated in the rumen (Conte et al., 2017). Several different biohydrogenation pathways have been proposed for LNA and ALA. Several factors including forage-to-concentrate ratio and ruminal passage rate/residence time can determine the pathway and extent of the biohydrogenation of LNA and ALA (Chilliard et al., 2014; Dugan et al., 2018; Dewanckele et al., 2020).

The main pathways for the biohydrogenation of LNA and ALA have been described by Harfoot and Hazlewood (1997). Pathways for both LNA and ALA are characterized by initial isomerization of the cis double bond at carbon 12 to a trans double bond at carbon 11 resulting in the production of cis9, trans11-18:2, an isomer of CLA (also known as rumenic acid; RA), and cis9, trans11, cis15-18:3, respectively ( Figure 1 ). This is followed by rounds of hydrogenation and isomerization leading to a trans 18:1 isomer (e.g., trans11-18:1) and eventually complete hydrogenation to 18:0 (stearic acid) as the end product (Dewanckele et al., 2020).

It is noteworthy that the abovementioned pathways were elucidated when greater forage-to-grain ratios (forage-based diets) were fed. When feeding grain-based diets (e.g., feedlot diets), isomerization of the cis 9 double bonds for LA shifts towards a trans double bond at carbon 10, with the same happening for ALA shifts, resulting in the production of trans10, cis12-18:2 and trans10, cis12, cis15-18:3, respectively (Alves and Bessa, 2014). These have been referred to as the “trans-10 shifted” biohydrogenation pathways ( Figure 1 ) which result in the accumulation of trans10-18:1 as the main TFA in ruminant products such as beef from feedlot (grain-finished) cattle (Alves and Bessa, 2014; Alves et al., 2021). Whereas trans11-18:1 (trans vaccenic acid; TVA) is typically the main TFA found in milk and meat from ruminants fed forage-based diets (e.g., grass-fed beef) (Jaakamo et al., 2019). Findings from animal model studies suggest that trans10-18:1 may harm health in a similar fashion to industrial TFA (Alves et al., 2021). On the other hand, TVA can be converted to RA (cis9, trans 11-CLA), both of which have been associated with several health benefits such as prevention of cancer, cardiovascular disease, and inflammation as well as improved immune function (Derakhshande-Rishehri et al., 2014; Kim et al., 2016). Consequently, TVA and RA are sometimes referred to as “good” TFA (Diane et al., 2016; Vahmani et al., 2020).

In addition to the main biohydrogenation pathways mentioned above, there are numerous minor pathways active in the rumen resulting in a plethora of biohydrogenation intermediates including conjugated and non-conjugated trienoic (18:3), dienoic (18:2) and monoenoic (18:1 and 16:1) TFA isomers (Vahmani et al., 2020). In fact, milk and meat fats from ruminants (e.g. cattle, sheep and goats) have the most complex fatty acid composition (> 100 different fatty acids) among all edible fats, in part due to the presence of numerous biohydrogenation intermediates. However, the human health effects of many of these intermediates are for the most part unknown.

The biohydrogenation process results in about 50 different types of TFA including trans-16:1 (trans6- to trans12-16:1), trans-18:1 (trans4- to trans16-18:1), conjugated 18:2 known as CLA (≥ 12 different CLA isomers with cis/trans or trans/cis configurations), non-conjugated non-methylene interrupted 18:2 known as atypical dienes (≥ 10 different isomers with cis/trans or trans/cis configurations) and conjugated 18:3 (≥ 3 different isomers with cis/trans/trans or cis/trans/cis configurations). Trans-18:1s are the predominant TFA (70-80% of total TFA), followed by CLA (10-25% of total TFA), atypical dienes (5-15% of total TFA), trans-16:1 (5-10% of total TFA) and conjugated linolenic acid (CLnA, <5% of total TFA) ( Table 1 ). Among individual TFA isomers, TVA, RA and trans9-16:1 (trans palmitoleic acid; TPA) have been the most studied isomers in terms of health effects and bioactivity, due to their high prevalence in ruminant foods and commercial availability (i.e. pure fatty acid isomers).

There have been numerous studies done on the health effects of industrial TFA during the past 4 decades. These studies have consistently found that industrial TFA have been associated with an increased risk of cardiovascular disease, mainly by lowering HDL and raising LDL levels (Stender et al., 2008; Brouwer et al., 2010; Radtke et al., 2017; Oteng et al., 2019). In the mid-1990s, many developed countries including the EU made recommendations to limit the intake of industrial TFA to 1% of the daily energy intake (Wanders et al., 2017). According to a study in 2010, the average daily TFA intake for adults was around 2.5 to 3.49% of daily energy intake in the U.S. (Micha et al., 2014), which was much higher than the 1% recommended limit. About 10 years later in 2015, the U.S. Food and Drug Administration (FDA) ruled that industrial TFA are not safe in food and set a June 2018 deadline for their removal from the food system (USDA, 2020). The official ban on industrial TFA has left ruminant-derived fats as the sole dietary source of TFA in the U.S., as well as in Canada and most European countries.

The estimated dietary intake of ruminant TFA varies between 0.8% to 1.7% of total energy intake depending on the country, with the average intake of ruminant TFA in the U.S. estimated to be about 1.2% of energy intake (Gebauer et al., 2011). In a more recent study, however, ruminant TFA intake in Europe and the U.S. was estimated to be around 0.5% of energy intake (Brouwer et al., 2013). Given recent bans on industrial TFA, there is need for new studies to determine the current intake of TFA from ruminant derived foods.

The TFA composition of ruminant derived foods is largely influenced by dietary, management and animal factors. Among these factors, diet composition is the main determinant of biohydrogenation pathways, and consequently of the content and composition of TFA in ruminant fats. Adding sources of PUFA to the diet (e.g. plant oil and oilseeds) significantly increases the contents of TFA in ruminant milk and meat including RA, TVA and TPA (Scollan et al., 2017; Guillocheau et al., 2020; Guo et al., 2023). The source of PUFA has the largest impact, with LNA-rich oils, yielding the greatest RA, TVA and TPA contents (Bessa et al., 2015; Kliem and Shingfield, 2016). Feeding ALA-rich sources also enhances CLnA content in ruminant fats (Kliem and Shingfield, 2016; Chikwanha et al., 2018). Seemingly effective novel alternative oil sources for increasing TFA, such as insect oils, warrant further investigation (Hervás et al., 2022). Furthermore, the amount of PUFA increases TFA content, reaching a peak when feeding between 50 and 80 g/kg DM intake (Scollan et al., 2017; Chikwanha et al., 2018). Besides PUFA, feeding forage-based diets, as opposed to grain-based diets, effectively increases TFA with trans-11 double bonds (TVA and RA) in ruminant fats (Chikwanha et al., 2018; Cabiddu et al., 2022). However, the effectiveness of forage-based diets at increasing TFA is determined by several factors relating to the source of forage including species diversity, cultivars, phenological stage, maturity, conservation method, particle length, presence of bioactive compounds and seasonality (Frutos et al., 2020; Cabiddu et al., 2022). Noteworthy, feeding a combination of forages and PUFA for an extend period substantially enhances presence of TFA in ruminant fats (Kliem and Shingfield, 2016; Vahmani et al., 2020; Alves et al., 2021), particularly when forage and PUFA sources are feed separately (Vahmani et al., 2017).

High-grain diets supplemented with LNA substantially increases the content of trans10-18:1 and trans10,cis12-CLA (i.e., trans10-shift) in ruminant meat and milk (Mapiye et al., 2015; Kliem and Shingfield, 2016). The t10-shift is exacerbated by feeding small grains (i.e. barley and wheat versus corn), pelleting, and increasing feeding duration (Mapiye et al., 2012; Mutsvangwa et al., 2012). A multitude of strategies to avoid the trans10-shift such as the addition of forages, non-starch fibers, strong buffers, antibiotics, antioxidants, yeast, chitosan, and agro-industrial by-products to high-grain diets, have been attempted with varying success (Alves et al., 2021; Amin and Mao, 2021; Hervás et al., 2022). Additionally, PUFA protective treatments and adsorbents inhibit the trans10-shift to a limited extent (Guo et al., 2023).

Notably, contents of TFA are more effectively increased in milk versus meat and in small versus large ruminants (Chilliard et al., 2007; Dugan et al., 2011; Chikwanha et al., 2018). The TFA contents in milk are somewhat influenced by animal individuality, breed, stage of lactation and parity (Samková et al., 2012). In meat animals, breed, sex, slaughter age and weight, anatomical location of fat depot and muscle type have marginal effects on TFA contents (Mapiye et al., 2015).

Most published data on TFA metabolism comes from studies on trans 18:1, which are the predominant fatty acid type in both ruminant and industrial TFA. Trans 18:1 isomers are intestinally absorbed to the same extent as cis 18:1 isomers and the double-bond position has little or no effect on their absorption (Baer et al., 2003). After absorption, trans 18:1 isomers can be incorporated into cell membrane phospholipids, or stored in the triacylglycerols of adipose tissues. Trans 18:1 isomers can be metabolized by oxidation, elongation, and desaturation processes, which result in isomer-specific metabolites with different biochemical properties (Vahmani et al., 2020). For TVA, two main metabolic fates have been characterized which include chain shortening to TPA and delta-9 desaturation to RA ( Figure 2 ). It has been estimated that ~19% of TVA, the main t-18:1 isomer in milk and meat from forage-fed ruminants consumed by humans, can be converted to RA by tissue-level Δ-9 desaturation which is catalyzed by the stearoyl-CoA desaturase 1 (SCD1) enzyme (Turpeinen et al., 2002; Miller et al., 2003).

In addition to Δ-9 desaturation, chain shortening of TVA by β-oxidation (peroxisomal β-oxidation of TVA) can lead to elevated levels of TPA in the plasma or tissues after consuming foods containing TVA, particularly grass-fed beef and dairy products. The conversion rate of TVA to TPA has been estimated to be 10% in cultured rat hepatocytes incubated with TVA (Jaudszus et al., 2014), however, the whole body (in vivo) conversion rate of TVA to TPA is not known. Given the very low concentration of TPA in the food supply including ruminant-derived foods (<0.05% of total fatty acids), the major origin of circulating TPA in humans is assumed to be from TVA intake from consumption of ruminant derived foods (Jaudszus et al., 2014).

The effects of ruminant- versus industrial- TFA on human health have been controversial and a subject of debate for many years. The recent removal of industrial TFA from the food supply in developed countries has renewed interest in understanding the human health effects of ruminant TFA. Several epidemiological studies have shown that, in contrast to industrial TFA, ruminant TFA do not appear to increase cardiovascular disease (CVD) risk and mortality (Ascherio et al., 1994; Pietinen et al., 1997; Jakobsen et al., 2008; Bendsen et al., 2011; Sacks et al., 2017). This has been attributed to the different isomeric profile of trans18:1 between ruminant and industrial TFA, as well as the low concentrations of total TFA in ruminant fats (2-6% of total fatty acids) compared to partially hydrogenated vegetable oils (60-65% of total fatty acids) (Stender et al., 2008). Conversely, based on recent human clinical trials, both industrial- and ruminant- TFA adversely affect cholesterol homeostasis (i.e. increased blood LDL-cholesterol and reduced HDL cholesterol) when consumed at comparable levels (Motard-Bélanger et al., 2008; Brouwer et al., 2013; Gebauer et al., 2015; Stender, 2015; Verneque et al., 2022). However, there is limited information available on the effect of ruminant TFA on other cardiovascular disease risk markers and the development of other chronic diseases such as type 2 diabetes. In the following section, we summarize published data from observational studies, clinical trials and animal studies on the effects of ruminant TFA on metabolic parameters related to type 2 diabetes.

Taken together, although TVA and its metabolites (TPA and RA) have been touted as having antidiabetic properties based on data from observational studies and a small number of animal studies, while the same effects have not been observed in other animal studies or in human clinical trials. Additional clinical and mechanistic studies are needed to better understand the isomer-specific effects of ruminant TFA. Nevertheless, based on the current knowledge regarding potential adverse effects of ruminant TFA on blood lipoprotein profiles, practices resulting in increased levels of this group of fatty acids in ruminant milk and meat should be carefully reconsidered.

PV: Conceptualization, Writing – original draft, Writing – review & editing. YX: Writing – original draft. MD: Writing – review & editing. CM: Writing – original draft, Writing – review & editing.