Historically, bile acids (BAs) were thought of as playing a role in lipid metabolism. In recent years, however, they have been found to affect the body in a much more wide-ranging way and are important in many normal functions and, therefore, also many disease states. CLI chatted to Dr Giovanny Rodriguez Blanco, lab leader at the Clinical Institute of Medical and Chemical Laboratory Diagnostics, Medical University of Graz, Graz, Austria, to find out more about the role of BAs in bone and muscle diseases like osteoporosis, osteosarcopenia and sarcopenia

Skeletal muscle atrophy (Adobe Stock)

Photomicrograph showing decreased fiber size with increased spacing between them, reduced myofibrils, increased endomysial connective tissue with fatty infiltration.

What are bile acids and what is their normal role in the body?

Bile acids (BAs) are steroid acids and are classified as primary or secondary BAs. Primary BAs are synthesized in the liver from cholesterol and stored in the gall bladder as an important component of bile. They mainly include cholic acid (CA), chenodeoxycholic acid (CDCA) and their glycine and taurine conjugated products (Fig. 1). Secondary BAs are synthesized by further reactions (deconjugation, dehydrogenation, and dihydroxylation) by the gut microbiota. BAs were originally considered as emulsifiers as they are released after a meal and help with lipid absorption. However, in the last few years, the understanding of BAs has changed significantly and now we think of them as hormones as they are capable of reaching virtually almost every organ and they can affect their metabolism. So, as such, BAs are now considered as signalling molecules and that’s the reason we are interested in them.

Why is measuring them necessary?

As a result of their wide involvement in normal physiology, BAs are also implicated as having an impact in a wide range of disease states when their levels are aberrant. As well as many of the metabolic diseases (such as type 2 diabetes, obesity, the various liver diseases, cholestasis of pregnancy, etc), BAs have been implicated in cardiovascular diseases, colorectal cancer, and diseases of the nervous system including Alzheimer’s disease. In our lab, in the metabolism area of the Clinical Institute of Medical and Chemical Laboratory Diagnostics at the Medical University of Graz, Graz, Austria (https://labordiagnostik.medunigraz.at/en/), we run BAs for diagnostics on a weekly basis and on top of that, in our research lab, we investigate their role in maintaining bone and muscle health. Altered levels of BAs correlate with diseases like osteoporosis, osteosarco-penia, and sarcopenia, and BA receptors have been found in both bone and muscle tissue. However, their function in keeping the skeleton healthy is not fully understood. In our lab, we investigate how BAs are modulating bone formation and resorption, as well as maintaining skeletal muscle mass and function, using preclinical models. Besides, based on our preliminary results in patients with osteosarcopenia, we hypothesise that they could be used as diagnostic or prognostic markers for musculoskeletal diseases.

Figure 1. Chemical structure of some common bile acids and the synthesis primary bile acid and generation of secondary bile acid

(a) initiation of synthesis by 7-hydroxylation of sterol precursors, (b) further modification of the ring structures, (c) oxidation and shortening of the side chain, (d1) hydrolyzation of the bile acids, (d2) conjugation of the bile acids with glycine, (d3) conjugation of the bile acids with taurine, (e1) dehydroxylation of the bile acids, (e2) conversion of chenodesoxycholic acid acid 7α-hydroxyl to 7β-hydroxyl.

Reproduced under the Creative Commons licence CC BY 4.0 (https://creativecommons.org/licenses/by/4.0/) from Liu J, Zhao Q, Qu C et al. Profiling bile acid composition in bile from mice of different ages and sexes. Front Physiol 2025;16:1626215 (https://doi.org/10.3389/fphys.2025.1626215). Copyright © 2022 Zhao, Liu, Sun, Yao, Yang and Wang.

How is analysis of bile acids normally achieved?

Over the years, a number of methods have been used to detect BAs, including enzyme analysis, radioimmunoassays, enzyme-linked immunosorbent assays (ELISAs), NMR and spectrophotometry. Generally, the main limitation of these methods is that they measure total BA and struggle to distinguish different BA species. Liquid chromatography mass spectrometry (LCMS), which is the method we use, has developed rapidly and become the mainstream method for BA detection as it can provide accurate quantitation of multiple species, including the primary, secondary and conjugated bile acids. Right now, we operate on a weekly basis and quantitate 15 BAs (Table 1), and we are working on increasing this number up to 60.

What is the potential for bile acid to serve as markers for bone and muscle disorders?

The gut microbiota plays an important role in metabolizing primary BAs into secondary BAs, some of which are important in muscle function and health and bone health. Bone and muscle disorders (such as osteoporosis, sarcopenia and osteosarcopenia) are generally related to ageing and, from data published on centenarians, we know that the gut microbiota is different in older people compared to younger people and that this changes the specific BA species that are produced, which alters the interaction of BAs with receptors in the bone and muscle. Besides, both primary and secondary BAs have been found altered in patients with these conditions. We, therefore, are interested in expanding the knowledge on BAs as endocrine regulators of bone and muscle by identifying novel BA species that might play a role in these tissues. In collaboration with some colleagues in Hungary, we profiled BAs in a group of post-menopausal women with low bone mass, sarcopenia, and osteosarcopenia, and found that the total primary BAs were reduced in osteosarcopenia, in contrast with an elevation of glucuronidated and sulphated BAs in sarcopenia compared to non-sarcopenia [Balasso et al., 2025].

This is our first investigation into the possibility of using BAs as a marker for these diseases. However, one of the limitations of the study is that our cohort was small, with 100 women in total, so more or less 30 per group, and so we need to expand the BA profiling in an extended and independent cohort to validate the findings. Currently the challenge is to find an extended cohort with such a good disease characterization, but recruitment is going well and we hope that next year we will have more patients to analyse.

Table 1. Bile acids (BAs) currently quantitated routinely by mass spectrometry in our laboratory

Are there any sex-related differences in bile acid levels?

Yes, we recently published another study where we analysed BAs in metabolic dysfunction-associated steatotic liver disease MASLD, formerly known as non-alcoholic fatty liver disease. We analysed a cohort of patients with MASLD and found an elevation of the classic bile acids in the cohort. When we disaggregated the data by sex, we noticed that the change was even higher in women than in men [Fitzinger et al., 2024]. So definitely biological sex has an effect on the BA levels. This is most likely due to the levels of estrogens and hormones in the women, and that deserves a lot of attention. On the same line, a recent publication using mice also found that BA composition markedly varies with age and sex [Liu et al., 2025].

Cholic acid is a primary bile acid synthesized in the liver from cholesterol (Adobe Stock)

If the sex-related differences in bile acid levels are due to hormones, do those differences start to decrease after the menopause?

It’s a good question. Although we might think that menopause would balance the BA profiles between sexes, preclinical research has shown that differences are still present between males and females at advanced age, long past the fertility period [Liu et al., 2025]. We need to take into account that many more factors apart from estrogens, are involved in the BA metabolism, including the complex role of microbiota. Thus, both preclinical and clinical investigations are still required to address this question.

What do you envisage in the future for analysis of bile acids?

I think the future of bile acid analysis relies on both the accurate identification and quantitation of additional bile acid species. As I mentioned before, we currently focus on 15 BA species although we can now analyse up to 60, but there are many more that we could detect in different sample types. However, this not easy and then the analysis should also be accompanied by standard reference materials and round-robin tests to see that the analysis is accurate – reproducibly and precise – maybe also the implementation of in vitro diagnostic regulation (IVDR) approved kits, as IVDR compatibility is required now. It is very likely that some companies are currently developing such kits for BA analysis by LCMS, which could be very welcomed by the community. Additionally, we are also interested in the potential for high throughput measurements, meaning that with technological development, we could to quantify BA concentrations in less than 5 minutes, where right now it takes 20–25 minutes. Therefore, by reducing the turnaround time by 80% we would have the flexibility to measure more samples and to reach to more people. So in summary, in future I expect we will expand the quantitation of BAs by identifying more species and faster.

Clinically, now, we understand that BAs reach almost every organ in the human body. This makes them very attractive to study, as they are involved in multiple diseases, including cardiovascular, musculoskeletal, and metabolic diseases (including insulin resistance), as well as cancer and Alzheimer’s disease. So in future, I would expect to see better understanding of how BAs are involved in disease and that we can use them as much more widely as diagnostic and prognostic markers in many more diseases.

Sarcopenia is age-related muscle loss (Adobe Stock)

 Bibliography
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2. Fitzinger J, Rodriguez-Blanco G, Herrmann M, Borenich A, Stauber R, Aigner E, Mangge H. Gender-specific bile acid profiles in non-alcoholic fatty liver disease. Nutrients 2024;16(2):250 (https://doi.org/10.3390/nu16020250).
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The interviewee

Giovanny Rodriguez Blanco PhD, Senior Scientist, Head of the Special Analytics Area

Clinical Institute of Medical and Chemical Laboratory Diagnostics, Medical University of Graz, Graz, Austria

Email address: g.blanco@medunigraz.at