Research article · 2023 · Full text
International Journal of Advanced Biochemistry ResearchThe level of isoflavones (genistein and daidzein) in different chicken feed formulations
Volume 7(2) · Pages 43–46 · DOI: 10.33545/26174693.2023.v7.i2a.241 ↗
Research areasPhytoestrogen effects and researchDiet, Metabolism, and Disease
Abstract
Author-writtenThe objective of the present study was to detect and quantify the level of isoflavones (genistein and daidzein) in different chicken feeds which were formulated based on BIS standards 2007. Six samples of chick mash, chicken grower and chicken breeder feeds used for analysis were procured from poultry feed mills and the samples were incubated with β-glucuronidase for better detection of different isoflavone aglycones and metabolites. The extracted isoflavones were purified by C18 solid phase extraction and analysed by high performance liquid chromatography (HPLC). The mobile phase composition, gradient elution programme and injected volume were optimised for separation efficiency and sensitivity. It is an accurate and precise analytical methodology which shows good reproducibility and can be easily adapted to detect isoflavones.The average genistein concentrations were 279.85 ± 1.30, 84.48±1.97 and 148.26 ± 1.64 µg/100 g in chick mash, chicken grower and chicken breeder respectively. However, the average daidzein concentration was in 276.29 ± 1.57 µg/100 g chick mash, 51.67±1.11 g/100 g in chick grower and 153.16 ± 1.21 µg/100 g in chick breeder. The findings indicated that the concentration of genistein and daidzein in the feeds was positively correlated with the level of soybean in the feed since it is the predominant source of isoflavones.
Summary
Source-checkedPlain-language reading of the full paper, prepared with AI and checked against the text. Each point links to the authors' own words.
Why it matters
- Isoflavones are powerful phytoestrogens known for their ability to modulate endocrine functions and provide health benefits. View evidence ↗
- Because soybean meal is a primary component of livestock feed, isoflavones can transfer from the diet into animal products. View evidence ↗
- Isoflavones exist as glycosides with low activity but are converted by intestinal microflora into aglycones, which can then trigger an estrogenic stimulus. View evidence ↗
The question
- The study aimed to detect and measure the levels of genistein and daidzein in various chicken feed formulations based on 2007 BIS standards. View evidence ↗
What they did
- The researchers analyzed six samples each of chick mash, chicken grower, and chicken breeder feeds obtained from poultry feed mills. View evidence ↗
- Samples were treated with β-glucuronidase to detect aglycones and metabolites, purified using C18 solid phase extraction, and analyzed via high performance liquid chromatography (HPLC). View evidence ↗
- The average soybean meal (SBM) content in the feeds was 30% for chick mash, 13% for chicken grower, and 20% for chicken breeder. View evidence ↗
- UV-VIS detection was performed at 262 nm for genistein and 260 nm for daidzein. View evidence ↗
What they found
- The differences in genistein and daidzein content between the three feed groups were statistically significant (p ≤ 0.05). View evidence ↗
- While the correlation between soybean meal (SBM) levels and genistein was not significant, there was a significant positive correlation between SBM levels and daidzein content. View evidence ↗
What it means
- The authors concluded that isoflavone levels in chicken feed are positively correlated with the amount of soybean meal used, with chick mash containing the highest levels. View evidence ↗
Visual summary
1 visualsIllustrations explain the paper; they do not show study data.
Soybean Meal and Isoflavone Correlation
AI-generated conceptual illustration (broiler chicken feed containing soybean meal). It explains the idea and does not show study data.
View source evidence ↗Figures & tables
From the original paper
Table 1
Figure 1
Table 2
Figure 2
Figure 3
Figure 4
Table 3Introduction
Isoflavones are potent phytoestrogens which have been widely studied because of their health benefits in modulating endocrine functions. Soybean meal constitutes a main ingredient of livestock feed formulations and soy isoflavones content have been evaluated in animal products due to their transfer from the diet. Isoflavones reside as glycosides with low estrogenic activity compared to their deglycosylated aglycone form. Upon ingestion, these compounds are metabolically hydrolysed by the intestinal microflora to their aglycones thus potentially mediating an estrogenic stimulus. Chick mash, chicken grower and chicken breeder feeds are types of specialized poultry feeds designed to meet the nutritional needs of chickens at different stages of their life cycle. These feeds are formulated to provide the appropriate balance of nutrients for chicks, growing chickens and breeding chickens to support their growth, health and reproduction.
Materials and Methods
Materials and Methods
Experimental diet
Chick mash is a type of starter feed designed for newly hatched chicks up to approximately 8 weeks of age. It is typically in the form of finely ground pellets or crumbles to make it easy for young chicks to consume. Chicken grower feed is intended for chickens that have outgrown the chick stage but are not yet ready for breeding or laying eggs. It is suitable for chickens from 9 to 16 weeks of age. Chicken breeder feeds are formulated specifically for mature chickens that are used for breeding purposes, whether for meat production or egg laying. The average incorporation level of soybean meal (SBM) was 30, 13 and 20 percent respectively in chick mash, chicken grower and chicken breeder feeds. The feed formulation of chick mash (CM), chicken grower (CG) and chicken breeder (CB) are enlisted in table 1.
| Ingredients | CM | CG | CB |
|---|---|---|---|
| Maize | 48.2 | 52.4 | 53 |
| Dorb | 10 | 22.7 | 15.5 |
| SBM | 29.9 | 13.1 | 20.3 |
| Cacite | 2.2 | 2.2 | 4 |
| Grit | 0 | 0 | 5 |
| Dcp | 1.5 | 1.6 | 1.3 |
| Methionine | 0.2 | 0.2 | 0.2 |
| Lysine | 0.1 | 0.4 | 0.2 |
| Soda Bicarb | 0 | 0 | 0.15 |
| Salt | 0.5 | 0.5 | 0.4 |
| Wheat Bran | 7.5 | 7 | |
| Total | 100 | 100 | 100 |
| Feed Additives | |||
| Vitamin Premix | 50 | 50 | 75 |
| Toxin Binder | 100 | 100 | 100 |
| Livertonic | 30 | 30 | 25 |
| Natchol | 50 | 100 | 200 |
| Tmmix | 125 | 10 | 150 |
| Avizyme | 35 | 35 | 35 |
| Coccistac | 50 | 50 | 0 |
Table text extracted from the original PDF; superscript letters are shown inline. The original table image is below.
View original table image
Standards and mobile phase for HPLC
All the chemicals and reagents of analytical grade were procured from M/s Sigma Aldrich India Pvt. Ltd. and M/s Merck Specialities Pvt. Ltd., Banglore. Stock solutions of free isoflavones (genistein and daidzein) were prepared in volumetric flasks by dissolving 2 mg of standard in 2 mL of HPLC-MS grade methanol (99.9%). Quantification and recovery tests were carried out with composite working standard solutions of 1 to 500 μg/mL which were prepared by diluting the stock solution with suitable quantities of methanol (Hu et al., 2019) [6] . Two different gradient elution systems were used as mobile phases to separate genistein and daidzein from samples (Saitoh et al., 2004) [8] . The gradients included an aqueous solvent A (0.1% acetic acid, 5% acetonitrile in water) and an organic solvent B (0.1% acetic acid in acetonitrile).
Feed sample preparation to estimate genistein and daidzein content
One gram of feed was weighed and extracted with 10 mL of 80 percent methanol. The samples were placed on an incubator orbital shaker for 1 h. The mixture was centrifuged twice @ 4500 rpm in high speed refrigerated microcentrifuge for 20 min. Extracts were mixed with 10mL of sodium acetate and 75 μL of β-glucuronidase. The solution was incubated overnight at 37 ºC and finally passed through C-18 purification kit before analysis.
Chromatographic and mass spectrometric conditions
Standard chromatogram and mass spectra were generated for each injected standard and test sample using DAD of chromatograph with a time window of 28 min. The column temperature was held constant at 35 °C. Twenty microliters of blank (mobile phase), samples prepared from control birds, standards and test samples were injected separately into the C18 column (3μm, 2.1X 150 mm) using an autosampler. The genistein and daidzein were separated by reverse phase column elution using a gradient program @ flow rate of 1 mL/min. The UV-VIS detection was carried out at 262 nm and 260 nm wavelength which was specific for genistein and daidzein.
Results
Chromatogram of genistein and daidzein standards are shown in figure 1. The height and area of the peaks were found to be increased with increasing concentration of standards. Calibration curve with the standards 1.95, 3.90, 7.81, 15.625, 31.25, 62.5, 125, 250 and 500 μg/mL were constructed for estimation of isoflavones. The retention time (RT) of standards was found to be in and around 18.99 to 19.14 min. for genistein, 12.24 to 12.28 min. for daidzein. Genistein and daidzein content of feed samples were significantly different (p≤ 0.05) among the groups. Higher mean values of genistein and daidzein were detected in chick mash (Fig. 2) feed samples followed by chicken breeder (Fig. 3) feed samples. On the other hand, chicken grower (Fig. 4) feed samples showed lower mean value for both genistein and daidzein. The table below (Table 2) presents the findings regarding the impact of soybean meal supplementation on the isoflavone content of feed samples. A non-significant positive correlation was observed between the SBM level and genistein content of feeds. However, it was significant and positively correlated between SBM and daidzein (Table 3).
| SBM / Isoflavone | SBM | — |
|---|---|---|
| Correlation coefficient | p-Value | |
| Genistein | 0.981 | 0.126 |
| Daidzein | 0.998 | 0.035 |
Table text extracted from the original PDF; superscript letters are shown inline. The original table image is below.
View original table image
| Groups | Mean value (µg/100 g) in feed samples | — |
|---|---|---|
| Genistein | Daidzein | |
| Chick Mash | 279.85a ± 1.30 | 276.29a ± 1.57 |
| Chicken Grower | 84.48c ± 1.97 | 51.67c ± 1.11 |
| Chicken Breeder | 148.26b ± 1.64 | 153.16b ± 1.21 |
| p-Value | ≤ 0.05 | ≤ 0.05 |
Table text extracted from the original PDF; superscript letters are shown inline. The original table image is below.
View original table image
Discussions
Soybean meal is a source of unsaturated fatty acids for energy utilization in livestock, especially in the poultry feed industry. Soy products are commonly used as animal feed due to their high protein content of about 38%, healthful fat and high fibre. This was the major driving force that stimulated the production and consumption of soybeans all over the world. The accuracy of identifying and quantifying isoflavones in feed samples were greatly influenced by the specific extraction and purification procedures and conditions (Gilani and Anderson, 2002) [4] . The most prevalent dietary isoflavones include genistein, daidzein, glycitein, formononetin, and biochanin A. These compounds are primarily found in legumes where they occur as glucosides (Liggins et al., 2002) [7] . Generally, soybean meal provided approximately 150 μg daidzein per gram and 250 μg genistein per gram (Dixon and Ferreira, 2002) [3] .
As per the review of Grgic et al. (2021) [5] , isoflavones were detected in 43% of cattle feed samples collected from January 2019 to April 2021 (n = 542) and the prevalence of the individual isoflavone in cattle feed ranged from 2.6% to 31%. Isoflavones were also detected in 45% of the pig feed samples, with daidzin and genistin as well as their aglycones being the most prevalent ones. The detected maximum concentrations were 140.0 mg/kg for genistein. Grgic et al. (2021) [5] also detected the presence of soy isoflavones in 88% of the poultry feed samples (n=263). Aglycones were detected in lower concentrations compared to the respective glycosides. The maximum concentrations were 102 mg/kg for genistein and 465 mg/kg for its glucoside genistin. The corresponding values for daidzein and its glucoside daidzin were similar but somewhat lower compared to genistein and genistin. Formononetin and biochanin A showed only low prevalence in poultry feed (0.8 and 5%, respectively). Ciftci (2017) [11] noticed that genistin and daidzin remained stable even when subjected to high temperatures during soybean processing and they were not lost during the traditional soybean processing methods. As a result, when domesticated poultry were fed with diets containing corn and soy led to complete absorption of all the genistin and daidzin present in soybeans which might be a contributing factor for higher isoflavone content in the different chicken feeds.
The current study findings were also supported by Silva and Perrone (2015) [9] who reported that high moisture thermal processing of soybeans being a procedure that was employed to improve the functional value of the product as it assumed to yield meals with potentially higher isoflavones bioavailability.
Soybean is mentioned to be one of the most important cultivated legumes throughout Asia Soy supplements are the richest source of isoflavones (Boettger-Tong et al., 1998) [1] with concentrations ranging from 118 mg/100 g to 306 mg/100 g. Therefore, soy foods and some foods with soybean additives had the largest concentration of these estrogenic compounds and were marketed because of their cost effectiveness (Vargas Galdos, 2009) [10] .
Conclusions
The free isoflavones such as genistein and daidzein were identified in different chicken feed formulations. The significantly higher mean values of genistein and daidzein were detected in chick mash feed followed by chicken breeder and grower feeds and it was positively correlated with the level of soybean meal incorporated in the feed.
Acknowledgement
AcknowledgementThe authors are thankful to Central Instruments laboratory, Kerala Veterinary and Animal Sciences University for providing the facilities needed for carrying out the research.
References
9 in the original paper- Boettger-Tong H, Murthy L, Chiappetta C, Kirkland JL, Goodwin B, Adlercreutz H, et al. A case of a laboratory animal feed with high estrogenic activity and its impact on in vivo responses to exogenously administered estrogens. Environ. Health Perspect. 1998 Jul;106(7):369-373.
- Sahu NK, Khajanji SN, Lakpale R. Effect of new premix herbicides molecules on yield attributing character and yield of soybean [Glycine max (L.) Merrill] in vertisols. Int. J Adv. Chem. Res. 2022;4(2):199-202. DOI: 10.33545/26646781.2022.v4.i2c.101
- Dixon RA, Ferreira D. Genistein. Phytochemistry. 2002 Jun 1;60(3):205-11.
- Gilani GS, Anderson JJ. Editors. Phytoestrogens and health. The American Oil Chemists Society; 2002 Jun 30.
- Grgic D, Varga E, Novak B, Müller A, Marko D. Isoflavones in animals: Metabolism and effects in livestock and occurrence in feed. Toxins. 2021 Nov 24;13(12):836.
- Hu L, Jin K, Zheng B, Yang X, Lei S. Simultaneous Determination of Isoflavones and Equol in Egg Yolk Using UPLC-MS/MS. Food Anal. Methods. 2019 Apr 15;12: 859-868.
- Liggins J, Mulligan A, Runswick S, Bingham SA. Daidzein and genistein content of cereals. Eur. J Clin. Nutr. 2002 Oct;56(10):961-6.
- Saitoh S, Sato T, Harada H, Matsuda T. Biotransformation of soy isoflavone-glycosides in laying hens: intestinal absorption and preferential accumulation into egg yolk of equal, a more estrogenic metabolite of daidzein. Biochim. Biophys. Acta BBA - Gen. Subj. 2004 Sep 24;1674(2):122-30.
- Silva F de O, Perrone D. Characterization and stability of bioactive compounds from soybean meal. LWTFood Sci. Technol. 2015 Oct 1;63(2):992-1000.
Cited by 10
From OpenAlex, OpenCitations and reference lists read by Scinovex
- Comparative effects of Thymus vulgaris supplementation on growth performance, hepatic enzyme activity, and antioxidant status in broiler chickens ↗
Ghada Ameen, Tahere Sarboozi Hosein Abadi, Mohammed M. Hassoon · Acta Entomology and Zoology · 2025
This study evaluated the effects of Thymus vulgaris supplementation on growth performance, hepatic biochemical indices, and antioxidant status in Ross 308 broiler chickens. A total of 120 one-day-old chicks (40±2 g) were randomly assigned to four treatment groups (n = 30 per group): a control group receiving a basal diet, a group supplemented with thyme powder (0.5 g/kg feed; T1),
- Comparative effects of Nigella sativa and Ocimum basilicum supplementation on growth performance, hepatic enzymes, and antioxidant status in broiler chickens ↗
Gheyath Hadi Ameen, Tahere Sarboozi Hosein Abadi, Mansoor Hasan Hassoon · Zoological and Entomological Letters · 2025
This study investigated the comparative effects of Nigella sativa seed powder and Ocimum basilicum aqueous extract individually and in combination on growth performance, hepatic enzyme activity, and antioxidant status in broiler chickens. A total of 120 one-day-old Ross 308 chicks (40±2 g) were randomly assigned to four groups (30 birds each): a control group fed a basal diet, a group
- Impact of alternative protein sources on growth and feed efficiency in broiler birds: A review
K. G. Vijay Anand · International Journal of Agriculture and Food Science · 2025
Broiler production relies on high-quality protein sources to support rapid growth, efficient feed utilisation and optimal carcass yield. Soybean meal and fish meal have long dominated commercial diets due to their balanced amino acid profiles and digestibility; however, rising global demand, fluctuating prices, land-use limitations and environmental concerns have accelerated the search for sustainable alternatives. This review synthesizes verified research on insect meals, fermented plant proteins, leaf meals, single-cell proteins and agro-industrial by-products as emerging protein replacement
- How different processing methods (roasting/extrusion/pelleting) change genistein and daizein levels in chicken feed ingredients ↗
Aisyah Nordin, Mohd Farhan Iskandar · Journal of Current Research in Food Science · 2025
Processing conditions applied during poultry feed manufacture can markedly influence the stability and bioavailability of soy-derived isoflavones, particularly genistein and daidzein, which are increasingly investigated for their physiological effects in poultry nutrition. The present study evaluates how three common industrial processing methods roasting, extrusion, and pelleting alter genistein and daidzein concentrations in selected chicken feed ingredients. Representative soybean-based ingredients were
- Low-cost IoT prototype for feed batch identification and isoflavone-level logging using QR codes and cloud sync ↗
Anurak Sittipornchai, Kanyarat Phromsuwan, Nattapong Rattanakorn · International Journal of Circuit Computing and Networking · 2025
Rapid diversification of poultry feed formulations has increased the need for reliable batch-level identification and compositional tracking to support quality assurance, traceability, and downstream research on bioactive compounds. Isoflavones such as genistein and daidzein are increasingly incorporated into commercial and experimental chicken feeds due to their reported biological relevance, yet their documentation at the batch level remains largely manual, fragmented,
- Evaluation of the performance of dinitroaniline herbicide on growth and yield of soybeans Glycine max L when different planting dates after spraying
Jasim Abdullah Hayawi, Raad Ahmed Hameed · International Journal of Agriculture and Nutrition · 2024
The research was conducted during 2022-2023 at the Rashedia Research Station in order to evaluate performance of the dinitroaniline herbicide when the Glycine max L. crop planting dates differed. The research encompassed two factors, the use of dinitroaniline herbicide is discussed at two levels, second: Planting date of the crop after spraying the herbicide is determined at three levels: immediate, a week, and two weeks post-spray. Research was applied using the method of the study utilized global experiments and a Randomized Complete Block Design (RCBD) with three replications. The data was
- Impact of algal-derived functional foods on animal health and performance ↗
Nathan Harris · Journal of Entomology and Zoology Studies · 2024
Algal-derived functional foods, particularly Spirulina (Arthrospira platensis), are gaining global recognition in animal nutrition for their dual role in providing high-quality nutrients and enhancing physiological functions. Rich in protein (55-70%), carotenoids, ?-linolenic acid (GLA), phycocyanin, vitamins, and minerals, Spirulina contributes to antioxidant defense, immune modulation, pigmentation, and gut stability. Evidence demonstrates improved growth, feed efficiency, and product quality in poultry,
- Feed-to-food transfer: A simple research proposal on isoflavone residues in chicken meat/eggs after feeding genistein-daidzein formulations ↗
Mariana Alves Ribeiro · International Journal of Pharmacology and Pharmaceutical Research · 2024
Isoflavones such as genistein and daidzein are naturally occurring phytoestrogens widely present in soybean-based feed ingredients and are increasingly explored for their functional roles in poultry nutrition. Their documented antioxidant, immunomodulatory, and growth-modulating properties have encouraged controlled inclusion of isoflavone-enriched formulations in poultry diets. However, the potential transfer of these bioactive compounds from feed into edible poultry products, particularly meat
- Effect of fermentation (Lactobacillus-based) on genistein and daidzein levels in soy-based chicken feed ↗
Liang Wenhao, Chen Ruiying · Journal of Advances in Microbiology Research · 2024
Soy-based feed ingredients are widely incorporated into poultry diets due to their high protein content and functional bioactive compounds, particularly isoflavones such as genistein and daidzein. These phytoestrogens are associated with antioxidant activity, modulation of lipid metabolism, and potential improvement in poultry health and productivity. However, the bioavailability of isoflavones in conventional soy-based feeds is limited because a substantial proportion
- Rapid UV-Vis / HPLC method optimization for quantifying genistein and daidzein in mixed chicken feed ↗
Lucas Henrique Moreira, Rafael Augusto Teixeira · International Journal of Chemical Research and Development · 2024
Isoflavones such as genistein and daidzein are increasingly incorporated into poultry feed formulations due to their phytoestrogenic, antioxidant, and growth-modulating properties. Accurate and rapid quantification of these compounds in complex feed matrices is essential for quality control, regulatory compliance, and feed-to-food safety evaluations. However, mixed chicken feed presents analytical challenges because of heterogeneous ingredients, variable fat and protein content, and