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. 2025 Aug 1;34(15):3645–3653. doi: 10.1007/s10068-025-01955-3

Antidiabetic effect of dried sweet potato extract with fermented lettuce extracts

Eunjin Kim 1,#, Soon Yeon Jeong 1,#, Ming Zhang 2,3, Yun-Seong Lee 4, Byeongjun Ji 5, Jungin Moon 5, Hyun-Ock Pae 4, Hyun Soo Chun 5,✉, Sooah Kim 2,✉
PMCID: PMC12528575  PMID: 41113272

Abstract

Fermented plant extracts are known to exhibit antidiabetic effects. However, antidiabetic activity of fermented lettuce extracts (FHE) and dried sweet potatoes with FHE remains unexplored. This study analyzed γ-aminobutyric acid (GABA), antioxidant, and polyphenol levels in such extracts, and an in vivo oral glucose tolerance, an oral sucrose tolerance, and a fasting blood glucose tests were conducted on streptozotocin-induced diabetes mice to determine antidiabetic effects. FHE showed higher GABA, antioxidant, and polyphenol levels than unfermented extract. The blood glucose levels in diabetic mice that were fed FHE and dried sweet potato with FHE were lower than those in mice that were fed distilled water. Mice fed dried sweet potato with FHE extract showed faster recovery and improved insulin sensitivity. These findings suggest that FHE has antidiabetic properties, which can be attributed to its high levels of GABA, antioxidants, and polyphenols, making it a potential candidate for antidiabetic food products.

Keywords: Fermented lettuce extract, Antidiabetic effect, Diabetes, Dried sweet potato

Introduction

Bioactive compounds have a wide array of health benefits because they exhibit antidiabetic, anti-inflammatory, antioxidant, antibacterial, antifungal, and antiviral properties (Boonphong et al., 2007; Combarros-Fuertes et al., 2019; Denaro et al., 2020; Skrovankova et al., 2015; Torres-Rego et al., 2016; Tran et al., 2020). Fermented plant extracts, which are functional foods, are rich in bioactive compounds. Fermentation induces structural changes that lead to the production of new bioactive compounds and extracts, which exhibit low cytotoxicity (Sadh et al., 2018). Fermentation can also increase the concentration of bioactive compounds (Limon et al., 2015; Punia et al., 2021).

Diabetes is a progressive metabolic disease characterized by chronic hyperglycemia resulting from insulin resistance and pancreatic β-cell dysfunction (American Diabetes Association, 2014). Current antidiabetic drugs are ineffective and have undesirable side effects; therefore, investigating new treatments is important (Yeap et al., 2012). Fermented plant extracts have received much attention as a new treatment option due to their antidiabetic properties. Fermented ginseng extract effectively controls Type 1 diabetes by inhibiting β-cell damage (Kim et al., 2010), and fermented mung bean extract reduces blood sugar levels by improving insulin secretion (Lim et al., 2012). Fermented Morinda citrifolia decreases serum insulin levels and insulin resistance (Jeon et al., 2013), and fermented soybean extract prevents hyperglycemia by reducing plasma glucose levels and increasing plasma insulin levels (Cheon et al., 2015). Although lettuce exhibits antidiabetic and anti-inflammatory properties (Kim et al., 2016), to our knowledge, there are no studies on the antidiabetic effects of fermented lettuce extract (FHE).

Sweet potato (Ipomoea batatas) is another functional food known for its high dietary fiber and bioactive phytochemicals content (Alam, 2021; Meng et al., 2019; Tian et al., 2016). However, the natural sugars in sweet potato may limit its effectiveness in diabetic management (Ooi and Loke, 2013). To overcome these limitations, we combined dried sweet potatoes with FHE, as they provide complementary bioactive compounds that may enhance overall antidiabetic effects. Therefore, we evaluated the antidiabetic properties of a combination of dried sweet potato and fermented lettuce extract in this study. Studies have shown that γ-aminobutyric acid (GABA), antioxidants, and polyphenols are responsible for the antidiabetic effects of fermented plant extracts (Sivamaruthi et al., 2018). Hence, we compared the amounts of GABA, antioxidants, and polyphenols in lettuce extract (LE) and FHE in vitro to investigate whether the antidiabetic effect of lettuce increases upon fermentation.

Materials and methods

FHE and dried sweet potato

HumanEnos Co., Ltd. (Wanju-gun, South Korea) prepared LE and FHE (Chun, 2020). Fresh lettuce was purchased from local markets, cleaned with ozonized water for 20 h, and dried for 1 day. Then, it was pulverized to less than 4 mm using a cutting mill (KM Tech, Icheon, South Korea). A sample of the pulverized lettuce was centrifuged at 12,000 rpm for 10 min, and the resulting supernatant was used as LE. Bacillus subtilis (KCTC 1201BP) was cultured with the remaining pulverized lettuce in distilled water (1:9, w/v) at 37 °C for 15–25 days to promote fermentation. Thereafter, the supernatant, acquired by ultrafiltration, was sterilized and used as FHE.

The dried sweet potato was purchased from Malrin Co., Ltd. (Wanju-gun, South Korea). Twenty grams of dried sweet potato were extracted with distilled water at a ratio of 10 times the weight (v/w) at 100 °C for 1 h to obtain sweet potato extract (IBE). The same method was used to process FHE; the two extracts were mixed to obtain sweet potato extract with fermented lettuce extract (IB + FHE).

High-performance liquid chromatography (HPLC) analysis of FHE

To investigate new bioactive compounds created by fermentation, LE and FHE were analyzed using HPLC. The extracts were vacuum-dried for 12 h using a vacuum concentrator (NB-503CIR, N-Biotek, Bucheon, Korea) and then suspended in 1 mL of 50% methanol (v/v). The solutions were filtered using 0.2 μm membrane filters (hydrophilic PTFE; Advantec MFS Inc., Dublin, CA, USA). Each sample (10 μL) was injected into an HPLC system equipped with an LC-20AD pump, an SPD-M20A diode array detector, a CTO-20A oven, a CBM-20A controller, and an SIL-20A autosampler (Shimadzu, Kyoto, Japan). The sample was then separated using a Symmetry C18 column (3.9 × 150 mm, 5 μm) at 25℃. Two mobile phases—water/acetonitrile (95:5, v/v) and 0.1% trifluoroacetic acid—were used to elute the sample isocratically at a flow rate of 0.5 mL/min for 60 min. The sample was analyzed using a UV–Vis detector at 215 nm.

GABA analysis

GABA analysis was performed as previously described (Lu et al., 2010). Briefly, 3 mL of extract was mixed with 1.5 mL of 0.5 M NaHCO3 and 0.5 mL of 0.715 mg/mL 1-fluoro-2,4-dinitrobenzene at 60 °C for 1 h. The solution was filtered using a 0.2 μm membrane filter (hydrophilic PTFE). The derivatives were analyzed using HPLC and eluted isocratically with 0.5% ammonium acetate solution and acetonitrile (85:15, v/v) at a flow rate of 1 mL/min for 20 min. A Symmetry C18 column (3.9 × 150 mm, 5 μm) at 30℃ was used for separation, and GABA concentration was determined using a UV/Vis detector at 360 nm. The retention time of the GABA standard was approximately 6.5 min, and the GABA in the samples was identified by comparing the retention time with that of the standard.

Ferric ion reducing antioxidant power (FRAP) assay

To compare the antioxidant activity of LE and FHE, FRAP assay was performed according to a previous study, with some modifications (Benzie and Strain, 1996). Briefly, FRAP reagent was prepared by mixing 25 mL of 300 mM acetate at pH 3.6, 2.5 mL of 10 mM TPTZ, and 2.5 mL of 20 mM FeCl3·6H2O. The reagent was then mixed with LE and FHE in a 1:9 ratio at 25 °C for 30 min. The absorbance of the mixtures was measured at 593 nm using a UV–Vis spectrophotometer (UV-1800, Shimadzu). Ascorbic acid was used as the standard, and the results were expressed as milligrams of ascorbic acid equivalents per milliliter of the extract (mg AAE/mL extract).

Total phenolic content

The Folin–Ciocalteu method was used to measure the total phenolic content of LE and FHE (Al-Farsi et al., 2005). Each extract (16 μL) was mixed with 60 μL of Folin–Ciocalteu reagent and incubated for 5 min; then, 60 μL of 60 g/L sodium carbonate solution was added. The mixture was incubated in the dark for 90 min, and its absorbance was measured using a UV spectrophotometer (UV-1800, Shimadzu) at 725 nm, using 50% methanol as the blank. The total phenolic content was expressed as milligrams of gallic acid equivalents per gram of dry weight (mg GAE/g dry weight).

Animal experiments and induction of diabetes

Animal experiments were approved by the Wonkwang University Animal Experimental Ethics Committee (Approval No. WKU21-44). C57BL/6 mice were obtained from Samtaco (Osan, South Korea) and acclimatized for 1 week in a controlled environment set to 23 ± 2 °C with a 12 h light–dark cycle.

The mice were fasted overnight, and diabetes was induced by a single intraperitoneal injection of streptozotocin (STZ; 120 mg/kg body weight) in 0.1 M citrate buffer (pH 4.5). After 2 weeks, the STZ-injected mice were fasted for 12 h, and blood glucose levels were measured using a portable blood glucose meter (Glucotrend, Roche, Germany). Diabetic mice with blood glucose levels > 200 mg/dL were selected for study.

Oral glucose tolerance test (OGTT) and oral sucrose tolerance test (OSTT)

Upon confirmation of diabetes, mice were randomly assigned to two groups. The control (CON) group (n = 5) was fed distilled water, whereas the experimental group (n = 5) was fed FHE, IBE, or IB + FHE. Thirty minutes after administering the extracts (200 μL/kg body weight), glucose or sucrose (2 g/kg body weight) was orally administered (Tables 1 and 2). Blood samples were collected from the tail veins of the mice 0, 15, 30, 60, and 120 min after glucose or sucrose administration, and blood glucose levels were measured using a blood glucose meter (Accu-Check®, Roche, Berlin, Germany).

Table 1.

Experimental design for oral glucose and sucrose tolerance tests on diabetic mouse model administered with fermented lettuce extract

(A)
Test type Group Material Dose (μL/kg) Material Dose (g/kg) n
Glucose tolerance CON Distilled water – Glucose 2 5
FHE Fermented lettuce extract 200 Glucose 2 5
Sucrose tolerance CON Distilled water – Sucrose 2 5
FHE Fermented lettuce extract 200 Sucrose 2 5
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CON control group; mice were administered distilled water, FHE fermented lettuce extract; mice were administered fermented lettuce extract

Table 2.

Experimental design for oral glucose tolerance, sucrose tolerance, and fasting blood glucose tests on diabetic mouse model administered dried sweet potato with fermented lettuce extract

Group Material Dose (mg/kg) Material Dose (g/kg) n
Test type CON Distilled water – Glucose 2 5
MET Metformin 200 Glucose 2 5
Glucose tolerance IBE Ipomoea batatas extract 300 Glucose 2 5
FHE Fermented Lactuca sativa extract 300 Glucose 2 5
IB + FHE Ipomoea batatas with fermented Lactuca sativa extract 300 Glucose 2 5
Sucrose tolerance CON Distilled water – Sucrose 2 5
AC Acarbose 10 Sucrose 2 5
IBE Ipomoea batatas extract 300 Sucrose 2 5
FHE Fermented Lactuca sativa extract 300 Sucrose 2 5
IB + FHE Ipomoea batatas with fermented Lactuca sativa extract 300 Sucrose 2 5
Fasting blood glucose CON Distilled water – Sucrose 2 5
AC Acarbose 10 Sucrose 2 5
IBE Ipomoea batatas extract 300 Sucrose 2 5
FHE Fermented Lactuca sativa extract 300 Sucrose 2 5
IB + FHE Ipomoea batatas with fermented Lactuca sativa extract 300 Sucrose 2 5
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CON control group; mice were administered distilled water, IBE dried sweet potato extract group; mice were administered dried sweet potato extract, FHE fermented lettuce extract group; mice were administered fermented lettuce extract, IB + FHE combination group; mice were administered a mixture of dried sweet potato extract and fermented lettuce extract

Fasting blood glucose test (FBGT) and insulin resistance

To evaluate the antidiabetic effect of IB + FHE, fasting blood glucose levels and insulin resistance were measured. For FBGT, samples were administered to each group, and glucose levels in tail vein blood were measured at 0, 15, 30, 60, and 120 min using a blood glucose meter (Table 2). Serum insulin levels were measured using a mouse insulin enzyme-linked immunosorbent assay kit (Shibayagi Co., Ltd., Gunma, Japan). The insulin resistance index (HOMA-IR) was calculated as follows:

HOMA-IR=fasting\; blood\; insulin(μU/mL)×fasting\; blood\; glucose(mmol/L)/22.5.

Statistical analysis

GABA concentration, antioxidant capacity, total polyphenol content, and blood glucose level were expressed as means ± standard deviations. To evaluate differences between groups, Student’s t-test was conducted using Statistica (version 7.1; StatSoft, Tulsa, OK), with p values < 0.05 being considered statistically significant.

Results and discussion

HPLC analysis of LE and FHE

Various unknown peaks at retention times of 1.6–3.4 min and 11.2 min were present in the chromatogram of FHE but not in that of LE (Fig. 1). These peaks were the result of structural changes in LE compounds induced by microbial metabolism during fermentation (Pham et al., 2019).

Fig. 1.

Fig. 1

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High-performance liquid chromatography chromatograms of (A) lettuce extract and (B) fermented lettuce extract

GABA analysis

Next, we investigated whether the unknown compounds in FHE induced physiological changes by evaluating the antidiabetic effect of FHE. GABA is released from cells to regulate insulin secretion (Bansal et al., 2011; Ngo and Vo, 2019; Purwana et al., 2014); hence, in the absence of or with a reduction in GABA levels, the risk of dysfunction and/or inflammation associated with diabetes increases (van Bussel et al., 2016). The GABA level in FHE (41.81 ± 0.80 μg/mL) was approximately 1.64 times higher than that in LE (31.13 ± 3.56 μg/mL) (Fig. 2A), indicating that bacteria produce GABA during LE fermentation (Sahab et al., 2020). The abundance of GABA and free amino acids in fermented mung bean enhanced its antidiabetic properties (Yeap et al., 2012). The high levels of GABA in FHE might induce a similar effect.

Fig. 2.

Fig. 2

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(A) γ-aminobutyric acid, (B) antioxidant activity, and (C) total polyphenol levels in lettuece extract (LE) and fermented lettuce extract (FHE). The data are expressed as means ± standard deviations. *p < 0.05, ***p < 0.001 for LE vs. FHE

Antioxidant capacity

Diabetes induces oxidative stress and causes various types of tissue damage (Kaneto et al., 1999). Antioxidants can protect pancreatic β-cells against glucose toxicity and oxidative stress under diabetic conditions (Rahimi et al., 2005), thus alleviating diabetes (Matough et al., 2012; Vincent et al., 2009).

The FRAP values of the extracts are shown in Fig. 2B. FHE reduced more ferric ions (0.677 ± 0.008 mg AAE/mL) than LE (0.561 ± 0.019 mg AAE/mL) (p < 0.05). This result indicates that the antioxidant capacity of lettuce increased upon fermentation, in agreement with reports on other fermented plant products. The antioxidant activity of noni fruit increased 1.2 times after a 40-day fermentation (Guo et al., 2020). After fermentation, the IC50 value of dandelion in a 2,2-diphenyl-1-picryl-hydrazyl radical scavenging assay decreased from 0.088 to 0.075 mg/mL (Liu et al., 2020). Therefore, the structural breakdown of plant cell walls due to fermentation leads to the synthesis or liberation of various antioxidants and improves antioxidant activity (Hur et al., 2014).

Total polyphenol analysis

The antioxidant activity of FHE was higher than that of LE. An increase in the content of phenolic compounds during fermentation improves antioxidant activity (Leonard et al., 2021). Phenolic compounds inhibit diabetes by regulating carbohydrate metabolism, improving glucose uptake, protecting pancreatic β-cells, enhancing insulin action, and controlling signaling pathways for cell homeostasis (Dias et al., 2017). To test whether phenolic compounds were responsible for the increased antioxidant capacity of FHE, its total polyphenol content was determined and compared with that of LE. The total phenolic acid content of LE increased 1.48 times (from 0.088 to 0.131 mg GAE/mL) after fermentation (Fig. 2C). In comparison, the total polyphenol content of a coffee extract increased 1.18 times, from 0.76 to 0.90 mg GAE/mL, after fermentation (Haile and Kang, 2019). Therefore, our results suggest that fermentation can improve the antidiabetic effect of LE.

In vivo blood glucose concentration for FHE using OGTT and OSTT

FHE exhibited a stronger in vitro antidiabetic capacity than LE. Therefore, its in vivo antidiabetic activity was investigated using OGTT and OSTT with five C57BL/6 mice each. The mean weights of the mice in the CON and experimental groups did not differ significantly. During OGTT, the weights of the mice in the CON and experimental groups were 37.98 ± 5.32 and 32.68 ± 3.31 g, respectively. The corresponding weights during OSTT were 33.88 ± 6.92 and 31.34 ± 1.97 g. The blood glucose level in mice that were fed distilled water was 503.8 mg/dL 30 min after glucose loading, which decreased to 325.2 mg/dL at 120 min post-loading. These levels were higher than the value observed at 0 h (190.2 mg/dL). However, when the mice were fed FHE, their blood glucose levels were 387.6 and 227.8 mg/dL, 30 and 120 min post-loading, respectively. In the OGTT experiments, the blood glucose levels of the experimental mice were lower than those of the CON mice over 2 h (p < 0.05) (Fig. 3A), indicating that FHE effectively suppressed increases in blood glucose levels over 2 h.

Fig. 3.

Fig. 3

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Blood glucose levels of mice administrated distilled water (CON) and fermented lettuce extract (FHE) in (A) oral glucose tolerance test (OGTT) and (B) oral sucrose tolerance test (OSTT). The data are expressed as means ± standard deviations. *p < 0.05, **p < 0.01 for CON vs. FHE

In the OSTT experiments, the blood glucose levels of CON mice increased from 198.0 to 444.8 mg/dL 30 min after the administration of sucrose and then decreased to 269.4 mg/dL at 120 min post-loading. These blood glucose levels were higher than those observed at 0 min (198.0 mg/dL). The blood glucose levels of the experimental mice increased from 200.2 to 352.8 mg/dL at 30 min post-loading and then decreased to 180.6 mg/dL 120 min after sucrose administration. The CON and experimental mice exhibited statistically significant differences over 2 h (p < 0.05) (Fig. 3B), indicating that FHE significantly suppressed the elevation in blood glucose levels after sucrose loading. These results agree with previous reports showing that fermented plant extracts exert antidiabetic effects by preventing β-cell damage, improving insulin secretion in plasma, and enhancing insulin sensitivity (Ahren et al., 2015; Cheon et al., 2015; Lim et al., 2012; Otto et al., 2016).

In vivo blood glucose concentration for IB + FHE using OGTT and OSTT

The OGTT results for IB + FHE are provided in Fig. 4A. All the samples effectively suppressed the increase in blood glucose levels. Thirty minutes after administration, blood glucose levels decreased by 14.7% in the IBE group, 32.5% in the FHE group, and 24.8% in the IB + FHE group, compared to that in the CON group (Fig. 4A). Similarly, the OSTT results (Fig. 4B) demonstrated a reduction in blood glucose levels following sample administration. After 30 min, the levels decreased by 16.8% in the IBE group, 34.3% in the FHE group, and 26.6% in the IB + FHE group, compared to that in the CON group.

Fig. 4.

Fig. 4

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Blood glucose levels of mice administrated distilled water (CON), metformin (MET), acarbose (AC), dried sweet potato extract (IBE), fermented lettuce extract (FHE), and a mixture of dried sweet potato extract and fermented lettuce extract (IB + FHE) in (A) oral glucose tolerance test, (B) oral sucrose tolerance test, and (C) fasting blood glucose test. (D) Insulin levels and (E) HOMA-IR values. The data are expressed as means ± standard deviations. * p < 0.05 vs. CON

Fasting blood glucose levels over a 2 h period are shown in Fig. 4C. Compared to those in the CON group, blood glucose levels were lower by 20.2% in the IBE group, 38.2% in the FHE group, and 30.6% in the IB + FHE group. The IB + FHE group had significantly lower serum insulin levels than the IBE group (Fig. 4D). Additionally, the HOMA-IR index, an indicator of insulin resistance, was lower for the IB + FHE group than for the CON group, suggesting an improvement in insulin sensitivity (Fig. 4E).

Consistent with these findings, noodles containing a fermented lettuce extract have shown a better antidiabetic effect than the control (Jeong et al., 2021). FHE may help regulate glucose metabolism by inhibiting digestive enzyme activity and enhancing insulin sensitivity, thereby preventing postprandial hyperglycemia. Therefore, FHE can be used as a functional additive in food products to prevent diabetes.

This study demonstrated the antidiabetic activity of FHE and IB + FHE. The levels of bioactive compounds that influence antidiabetic activity, such as GABA, antioxidants, and polyphenols, were higher in FHE than in LE. In addition, OGTT and OSTT assays indicated that the blood glucose levels of mice that were fed FHE and IB + FHE were lower than those of CON mice. These results suggest that FHE exhibits significant antidiabetic activity and could potentially be applied as an antidiabetic functional food.

Acknowledgements

The author(s) received no financial support for the research, authorship, and/or publication of this article.

Declarations

Conflict of interest

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Ethical approval

All animal experiments were conducted in accordance with the institutional guidelines and were approved by the Institutional Animal Care and Use Committee (IACUC) of Wonkwang University (Approval No. WKU21-44).

Footnotes

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Eunjin Kim and Soon Yeon Jeong have contributed equally to this work.

Contributor Information

Hyun Soo Chun, Email: 119bio@naver.com.

Sooah Kim, Email: skim366@jj.ac.kr.

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