How renal toxins respond to renal function deterioration and oral toxic adsorbent in pH‐controlled releasing capsule

The number of patients with chronic kidney disease (CKD) is increasing. Oral toxin adsorbents may provide some value. Several uremic toxins, including indoxyl sulfate (IS), p‐cresol (PCS), acrolein, per‐ and poly‐fluoroalkyl substances (PFAS), and inflammation markers (interleukin 6 [IL‐6] and tumor necrosis factor [TNF]‐alpha) have been shown to be related to CKD progression. A total of 81 patients taking oral activated charcoal toxin adsorbents (AC‐134), which were embedded in capsules that dissolved in the terminal ileum, three times a day for 1 month, were recruited. The renal function, hemoglobulin (Hb), inflammation markers, three PFAS (PFOA, PFOS, and PFNA), and acrolein were quantified. Compared with the baseline, an improved glomerular filtration rate (GFR) and significantly lower acrolein were noted. Furthermore, the CKD stage 4 and 5 group had significantly higher concentrations of IS, PCS, IL‐6, and TNF but lower levels of Hb and PFAS compared with the CKD Stage 3 group at baseline and after the intervention. Hb was increased only in the CKD Stage 3 group after the trial (p = .032). Acrolein did not differ between the different CKD stage groups. Patients with improved GFR (responders) (about 77%) and nonresponders had similar baseline GFR. Responders had higher acrolein and PFOA levels throughout the study and a more significant reduction in acrolein, indicating a better digestion function. Both the higher PFOA and lower acrolein may be related to improved eGFR (and possibly to improvements in proteinuria, which we did not measure. Proteinuria is associated with PFAS loss in the urine), AC‐134 showed the potential to improve the GFR and decrease acrolein, which might better indicate renal function change. Future studies are needed with longer follow‐ups.

recruited.The renal function, hemoglobulin (Hb), inflammation markers, three PFAS (PFOA, PFOS, and PFNA), and acrolein were quantified.Compared with the baseline, an improved glomerular filtration rate (GFR) and significantly lower acrolein were noted.Furthermore, the CKD stage 4 and 5 group had significantly higher concentrations of IS, PCS, IL-6, and TNF but lower levels of Hb and PFAS compared with the CKD Stage 3 group at baseline and after the intervention.Hb was increased only in the CKD Stage 3 group after the trial (p = .032).Acrolein did not differ between the different CKD stage groups.Patients with improved GFR (responders) (about 77%) and nonresponders had similar baseline GFR.Responders had higher acrolein and PFOA levels throughout the study and a more significant reduction in acrolein, indicating a better digestion function.Both the higher PFOA and lower acrolein may be related to improved eGFR (and possibly to improvements in proteinuria, which we did not measure.Proteinuria is associated with PFAS loss in the urine), AC-134 showed the potential to improve the GFR and decrease acrolein, which might better indicate renal function change.Future studies are needed with longer follow-ups.

| INTRODUCTION
The prevalence of chronic kidney disease (CKD), characterized by the progressive loss of renal function over time, is increasing worldwide. 1is alarming trend, primarily driven by the surge in concomitant conditions such as diabetes (DM) and hypertension, underscores the urgent need for effective therapeutic interventions and predictive markers to delay renal function decline in CKD patients. 2The renal function marker serum creatinine (Cr) is known to be biased and has much interference during measurements. 3In addition, blood urea nitrogen (BUN) and serum Cr are not sensitive markers for renal function changes, especially for patients with CKD.
Indoxyl sulfate (IS) and p-cresol (PCS) are commonly used uremic toxin markers. 4IS and PCS originate from bacterial protein fermentation in the large intestine. 5For example, colonic microbiota degrades tryptophan into indole, and further hydroxylation results in 3-hydroxy-indole, the majority of which is sulfonated to produce IS.However, whether these markers have a causal relationship with renal function deterioration is still under debate. 6ven these challenges, there is a pressing need to explore alternative approaches for predicting and preserving renal function in CKD patients.We propose the exploration of acrolein as a potential biomarker for renal function, especially for DM-related CKD.Acrolein has been identified as a uremic toxin. 7Acrolein is a highly electrophilic α, β-unsaturated aldehyde associated with human diseases.It can be formed externally by ingesting food prepared by overheating cooking oils or smoking. 8It can also be formed internally through the metabolism of polyamines or lipid peroxidation. 9Not only does acrolein produce oxidative stress and exacerbate the progression of CKD but it also poses a threat to cardiovascular function due to impaired endothelial function. 10Studies have suggested that acrolein may play a key role in the pathogenesis of CKD, making it a potentially robust predictor of renal function decline. 9However, toxins which are sensitive to changes in the glomerular filtration rate (GFR) remains unknown.
Furthermore, per-and poly-fluoroalkyl substances (PFAS) have been shown to be related to health issues and renal function deterioration.Some PFAS have been classified as persistent organic pollutants and have long half-lives in humans.PFAS are mainly consumed orally and excreted by the kidneys. 11Patients with early CKD may have higher PFAS due to accumulation compared with normal people.
But, serum PFAS may be decreased as in the late CKD (Stage 3b to stage 5 until there is anuria) or by any proteinuria, as proteinuria has been implicated in PFAS loss in the urine. 12And, PFAS correlates with cholesterol, poor physical health, and impaired liver function. 13ong PFAS, perfluorooctanesulfonic acid (PFOS) has the highest amount, followed by perfluorooctanoic acid (PFOA) and perfluorononanoic acid (PFNA).These are artificially manufactured substances, have a wide range of applications, such as surfactants. 14But, several PFAS are banned in some countries due to their toxic effects and high persistence. 15 addition to showing promise as better uremic markers, oral toxin adsorbents have shown some promise in mitigating the deleterious effects of accumulated toxins in CKD patients. 16,17Recent studies have been aimed toward the development of drugs that have the potential to preserve renal function.These studies represent a significant shift from traditional symptomatic treatments for patients not undergoing dialysis to interventions that aim to address the underlying progression of the disease. 18T-120 (KREMEZIN ® ) has been shown to prolong the time to the initiation of hemodialysis, slow the decline in the GFR, and decrease serum creatinine. 16AST-120 consists of oral, spherical carbon particles that adsorb uremic toxins and their precursors within the gastrointestinal tract, allowing them to be excreted in the feces.
AST-120 has been shown to be associated with a significant dosedependent reduction in serum IS levels and a decrease in uremiarelated malaise. 19Another bamboo charcoal adsorbent, CharXgen, has been shown to decrease toxin levels in an animal trial.However, data on human trials are limited. 17spite the potential benefits of these adsorbents, their adoption has been constrained by several limitations.Both activated carbon adsorbents are uncoated and travel approximately 6-7 m through the oral cavity, esophagus, stomach, and small intestine until they reach the colon.Food takes about 2-4 h in the stomach and 3-7 h in the small intestine before entering the large intestine. 20cause of this, the potential adsorption sites of the activated carbon may have been already covered by food particles, raising doubts on how much adsorption capacity remains once it reaches the large intestine, where the toxic waste concentration is supposed to be the highest along the digestive tract.Although these adsorbents are suggested to be taken at least 2 h after a meal to avoid interacting with food, this suggestion decreases compliance since CKD patients are usually already on a lot of medications due to multiple comorbidities. 21aditional indicators of uremic toxins, such as IS and PCS, have been shown to decrease after taking these oral adsorbents. 5However, it is unclear whether this is due to the removal of toxins produced by the fermentation of amino acids or because the amino acid substrates (such as indoles) have been removed before reaching the small intestine.Also, the absorption of small-molecule nutrients, such as amino acids, may be interfered with and lead to poor nutrition statuses for CKD patients, which can adversely affect CKD patients who are already prone to nutritional deficiencies. 22T-120 is restricted in its ability to adsorb certain particle sizes (from 100 to 10 000 Da) to avoid the possibility of interfering with the nutritional adsorption of nutrients, such as large peptides. 16However, it may also mean it cannot clear large-molecule toxins, especially protein-bound ones, and many large-sized toxin protein-bound complexes, such as PFAS and acrolein, have been shown to have adverse effects on kidney function. 9,13ditionally, the addition of activated carbon-based adsorbents can result in harder stool consistencies, leading to possible constipation.The risk of constipation, a common side effect, may adversely impact patient compliance and overall quality of life in CKD patients. 16Improving the risk of constipation is crucial to the treatment.Dietary fiber and probiotics could be a possible solution, and fiber intake has been shown to shift the gut microbiota toward reduced production of uremic toxins, mitigate metabolic acidosis, and slow CKD progression. 23is study tested the efficacy of a new activated charcoal oral adsorbent embedded in pH-controlled releasing capsules (AC-134), a patented acid-resistant plant-based capsule.Each capsule contains 250 mg of active charcoal powder with 200 mg of dietary fiber with probiotics (such as inulin and xylo-oligosaccharides) (Appendix in Data S1).The capsule has a three-layer coating, which disintegrates at a pH of 7 after 1 h, allowing the activated charcoal to act precisely on the back end of the ileum.We hope this oral adsorbent may provide better toxin clearance, especially for larger toxin protein-bound complexes, such as acrolein and PFAS. 11

| MATERIALS AND METHODS
An Institutional Review Board (IRB) and Ethics Committee approval was obtained before the trial.The study was conducted in full compliance with the Declaration of Helsinki.This study was approved by Taipei City Hospital (IRB: TCHIRB-11106016).Informed consent was obtained from all subjects involved in the study.Written informed consent was obtained from the patients for the publication of this paper.
This study tested whether this new form of pH control released capsules (AC-134) improved the GFR by decreasing serum Cr and BUN and whether the changes correlated with the uremic toxin levels (IS, PCS, PFAS, and acrolein) and inflammation markers (interleukin-6 [IL-6] and tumor necrosis factor [TNF]).Each capsule contains 250 mg of active charcoal powder with 200 mg of dietary fiber with probiotics.
Participants with impaired renal function ranging in age from 18 to 90 were included in this study.Specifically, individuals diagnosed with proteinuria or stage 3-5 CKD with a GFR lower than 60 were recruited.The intervention involved the administration of activated charcoal, originally intended to adsorb toxins by gastric lavage, encapsulated within a capsule (AC-134) designed to disintegrate in the terminal ileum.All participants were instructed to take two capsules three times a day for 1 month (30 days).All patients included have not been dialyzed before.
Exclusion criteria included patients with renal failure who had received a transplant or ever had hemodialysis or peritoneal dialysis, patients receiving chemotherapy for malignancy, patients who had a blood transfusion or admission in the past 2 weeks or intravenous medication, such as a lipid-based nutritional supplement, propofol, dopamine, methotrexate, fluorouracil, vancomycin, prednisolone, furosemide, or cyclosporine, and patients who were unwilling to sign the study agreement.
Before and after the intervention, several parameters were assessed, including BUN, Cr, estimated GFR, hemoglobulin (Hb), IS, PCS, IL-6, TNF, and three kinds of PFAS.The detection of acrolein in the plasma using a competitive enzyme-linked immunosorbent assay was described previously. 9The measurement of PFAS has been described in a previous study 3 (Appendix 2 and 3 in Data S1).
A responder was defined as having an increase in GFR after the intervention.Patients were divided into two subgroups, responders and nonresponders, and the factors mentioned above were compared.This study recruited 81 patients.
The PFAS level of gender difference was not noted in previous studies in elderly. 24,25So the participants were not stratified by sex.
Statistical analyses were performed using SPSS 19.0.Categorical variables are expressed as the number and percentage (n [%]).Continuous data with a normal distribution are presented as the mean accompanied by the standard deviation (SD) or the median and interquartile range if the data are not normally distributed.
Statistical analyses involved t-tests to compare the continuous clinical parameters between the subgroups.For categorical variables, chi-squared tests were employed.To assess the differences between the pre-and post-intervention results, paired t-tests (if normally distributed) or Wilcoxon signed-rank tests were employed based on the normality assumption of the data.If a 5% chance of a type I error (α = .05)was permitted, with a power of 90%, assuming the difference in values before and after the intervention was at least equal to its SD, then an adequate sample size was at least 42 patients.A total of 81 patients were enrolled in this study.A p-value less than .05was considered statistically significant.

| RESULTS
This study recruited 81 patients aged around 70 years old, and about 70% were male.Initially, 38 patients had a GFR between 30 and 60 mL/min (CKD Stage 3), and 43 were in CKD stages 4 and 5.There were 63 responders out of the total 81 patients (response rate of 77%).
After the intervention, significant decreases were noted in the BUN, Cr, and acrolein levels, with an increase in GFR in the CKD Stage 3 and CKD stage 4 and 5 groups.In subgroup analysis, the CKD stage 4 and 5 group had higher BUN, Cr, IS, PCS, IL-6, and TNF levels throughout the study.In contrast, lower Hb and lower PFAS (PFOA, PFOS, and PFNA) levels were noted in the CKD stage 4 and 5 group compared with the CKD Stage 3 group.Moreover, the Hb was significantly higher after intervention in the CKD Stage 3 group after the study.Tables 1 and 2 summarize these findings, and Figures 1   and 2 visually represent these findings.Table 1 also demonstrates that the inflammation markers (IL-6 and TNF) and uremic toxin markers (IS and PCS) did not exhibit significant changes in relation to the GFR, and these variables appeared to remain relatively stable.
Figure 3 shows the linear regression of Hb with the sum of all the PFAS, PFOA, PFOS, and PFNA (all p < .05,except for PFOS).
After the intervention, acrolein levels were lower, and GFRs were higher in all the patients (Figure 4A,B) and in both CKD groups compared with their baselines (Figure 4C,D).However, no significant difference in acrolein was noted between the different stages of CKD.
Patients with an increase in GFR after the intervention were defined as responders.The overall response rate was 77% (63/81).
Chi-square tests identified the relationships between patient responses and variables such as sex, age, GFR, and the presence or absence of DM (Table 3).These findings revealed no significant relationships between these variables.

T A B L E 1
The demographic data of the participants (gender and age) and the clinical parameters before and after the study of taking AC-134 for 1 month.In comparing the groups with and without GFR improvements (responders vs. nonresponders), nonresponders were slightly older ( p = .072)and had marginally higher levels of TNF (p = .065)at the baseline and after the intervention ( p = .056)(Table 4).However, these differences were not statistically significant.
The two groups had no significant differences in the baseline BUN and Cr levels while responders had a significantly larger decrease in BUN and Cr levels than the non-responders.(Table 4).In the responders' group, the pre-and post-intervention acrolein and PFOA levels were significantly higher (Figure 5A

| DISCUSSION
Table 1 provides data indicating that as the GFR increased after the intervention, there were notable decreases in BUN, Cr, and acrolein levels, which were statistically significant.This data suggest a potential relationship between the GFR and these variables.These results imply a noteworthy association between the decline in BUN and Cr and the elevation in the GFR, which might be attributed to the significant decrease in acrolein, an identified uremic toxic compound, which leads to the progression of CKD (Figures 1 and 2). 9 Contrary to our initial hypothesis, IS and PCS did not exhibit significant changes (Figure 3).This finding provokes a reconsideration of the toxicity associated with IS and PCS, suggesting that the deleterious effects of these amino acid metabolites might not be as sensitive markers for CKD as previously thought. 4I G U R E 1 The indoxyl sulfate (IS) and p-cresol (PCS) (A and B), and the inflammation markers (C and D) between chronic kidney disease (CKD) Stage 3 (n = 38) and CKD stage 4 and 5 (n = 43) before and after the intervention, all the markers were higher in the CKD stage 4 and 5 group throughout the study (all p < .05).
In a previous study, it was reported that AST-120 (KREMEZIN ® ) may decrease the levels of IS and PCS. 19Whether this result was caused by the amino acid substrate of IS and PCS being adsorbed or the removal of toxins is unclear.Further studies are needed to answer this hypothesis.
The inflammation markers IL-6 and TNF did not demonstrate significant changes, providing further complexity to understanding CKD.
These findings indicate that the GFR improvement noted in the study may not directly correlate with inflammation, although inflammation is an important factor that triggers CKD progression. 1,9The beneficial effects are most likely to result from the decrease in uremic toxins other than IS and PCS, 26,27 because we did not observe significant changes in IS and PCS within 30 days.
However, when the patients were separated into two groups according to GFR, patients with lower GFRs (CKD stage 4 and 5) had higher IS, PCS, and inflammation markers (Figure 1) and had lower levels of Hb and all three PFAS (Figure 2).Anemia progression as renal function deteriorates is no surprise; however, PFAS levels were lower in the poor renal function CKD stage 4 and 5 group, contrary to our expectation since some PFAS levels are much higher in hemodialysis patients compared with normal people in our previous study. 15likely explanation for all or some of the finding is that PFAS bind to proteins and already known to be associated with proteins such as albumin and hemoglobin.28 Albumin was not measured in this study.Had it been measured, we expect that it would show a similar association. Figure 3 shows the linear correlation of Hb with the sum of all three PFAS, PFOA, PFOS, and PFNA (Figure 3A-D).
PFAS are highly protein-bound toxins that decrease as renal function worsens due to malnutrition, hypoalbuminemia, or anemia.PFOS is found at the highest level of all PFAS and may act with serum proteins, as shown in a previous study. 11So, the correlation of Hb with PFOS is not as significant as the other two PFAS: PFOA and PFNA.
In addition, this study showed that Hb increased from 11.73 to 12.06 (g/dL) ( p = .054)in all patients (Table 1) and with significantly higher Hb levels from 13.39 to 13.80 (g/dL) (p = .032)in the CKD Stage 3 group after the intervention (Table 2 and Figure 2A).This study also showed that the decrease in acrolein may improve the erythropoietic effect in CKD Stage 3 patients, while patients with CKD stage 4 or 5 may only have a beneficial effect in GFR because their erythropoietin (EPO) levels may be too low to show improvements.The improvement of Hb in CKD Stage 3 is not an effect of the artificial EPO injection because extra-EPO is only allowed in patients whose GFR is lower than 15 mL/min in Taiwan. 29As the erythropoietic response is highly affected by the inflammation status, which is also higher in CKD stages 4 and 5 (Both IL-6 and TNF are significant predictors of erythropoiesis), 15 we concluded that inflammation may be a key factor for anemia in the group of CKD stage 4 and 5.However, the decrease in acrolein may still be beneficial for erythropoiesis since acrolein stimulates suicidal erythrocyte death or eryptosis. 30gure 4A,B shows that acrolein was decreased and the GFR was increased in all 81 patients.When divided by GFR, there was no significant difference in acrolein levels between the CKD Stage 3 group and the CKD stage 4 and 5 group.However, the acrolein level is not related to the renal function from CKD Stage 3 to stage 4 and 5 (Figure 4C,D).One explanation is that acrolein is a protein-bound toxin like PFAS (which may decrease in serum concentration as malnutrition and anemia progress) and is also a uremic toxin, which causes CKD progression (which may increase in concentration as renal function worsens and is not eliminated from the body). 9These two effects may offset each other.
To find those most suitable for the oral toxic adsorbent (AC-134), Table 3 compares different variables, such as sex, age, GFR, and DM, between the responders and nonresponders; however, no significant differences were noted during these comparisons.These results imply The linear regression of hemoglobin (Hb) to PFAS (all PFAS, PFOA, PFOS, and PFNA) at the baseline before the intervention (A-D) (n = 81) (all p < .05except for PFOS).PFAS, per-and poly-fluoroalkyl substance; PFOA, perfluorooctanoic acid; PFOS, perfluorooctanesulfonic acid; PFNA, perfluorononanoic acid.
that the capsule (ACT-134) works equally well regardless of sex, age, DM status, and CKD stage.
Table 4 shows that the nonresponder group consisted of slightly older individuals with slightly higher levels of TNF (although not significant, .05< p < .1).Age was identified as a factor associated with poor response, and elevated TNF levels indicated inflammation, which may contribute to accelerated aging. 22 the responders' group, the pre-and post-intervention acrolein and PFOA levels were significantly higher (Table 4 and Figure 5A,B) throughout the study, with a more considerable decrease in acrolein and increase in GFR compared with the nonresponders (Figure 5C,D).
The baseline GFR was around 25 mL/min, and the median increase of GFR was around 2-3 mL/min in the responders' group.
Before the intervention, there was no significant difference in the GFR between the two groups.However, after the intervention, the responders' group was characterized by lower BUN and Cr levels and exhibited a significantly greater increase in the GFR.The significantly higher levels of acrolein observed before and after the intervention and the substantial decrease in acrolein concentration may suggest improved food absorption and the efficient elimination of toxins in the responders' group (Table 3 and Figure 4).Intestinal function may be the key point for effective oral toxic adsorbent.PFOA is an artificially made material and cannot be endogenously produced.Higher acrolein and PFOA levels may indicate better digestive function and nutrition status.One reason why the responders' group had higher acrolein levels may be reverse epidemiology, which has been noted in dialysis patients. 31Dialysis patients with malnutrition have been shown to have lower lipid levels but higher vascular events.This paradox is well-known, and the survival disadvantages of malnutrition may have a major impact on mortality. 22,32 contrast, higher amino acid metabolites and waste concentrations might indicate better food intake in CKD patients.Hence, those with a better nutrition status are more likely to have higher acrolein levels due to better food intake.The previous study result has shown that acrolein levels were unchanged in different CKD stages, so patients with higher acrolein levels may not necessarily have poorer renal functions.
Another interesting finding is that PFOS is decreased in responder group but PFOA and PFNA are increased after intervention, although not significantly.(Table 4).
PFOS are more affected and tend to decrease if the enterohepatic recirculation is disrupted, compared with PFOA. 33It may be related to the active charcoal and fiber contained in AC-134 that disrupts the resorption of PFOS from the intestine and colon. 34mpared with the baseline, the average GFR of all patients is improved and we found that PFOA and PFNA are increased after our intervention.The possible explanation is the improvement of GFR is accompanied by the decrease of proteinuria and lower the loss of PFOA and PFNA through the urine. 12Thus the average levels of PFOA and PFNA are increased.
In short, better gastrointestinal tract motility and clearance of toxins with significantly decreased acrolein values after intervention may offset the disadvantages of higher baseline acrolein levels.Acrolein is not only a uremic toxin that causes damage to the heart, kidneys, and cells 35 but is a carcinogen that forms DNA adducts, which may induce lung cancer and respiratory diseases. 36Acrolein may also produce damage to the intestinal barrier by stress-mediated cell death and is highly associated with an increase in insulin resistance and DM.
In the realm of therapeutics for CKD, pH-controlled release capsules (AC-134) have demonstrated the ability to protect the absorptive capacity of the activated charcoal, thereby effectively adsorbing toxins in the body with an average response rate of about 77% (63/81) (Table 4).The pH-controlled release capsules (AC-134) have several advantages over other uremic toxin adsorbents.First, the capsule retains its adsorption ability before it is saturated with small food particles, such as amino acids and peptides, before the ileum.So, AC-134 will not interfere with nutrient absorption. 16As a result, AC-134 can be taken with or without a meal, and this convenience may increase compliance. 21Other toxin adsorbents are bare particles exposed to the digestive tract and cannot completely avoid the adsorption of small nutrient elements. 17cond, the capsules ensure that this absorption extends not just to small molecules but also to larger substances bound to proteins, which are hard to remove.This broad-spectrum absorption can potentially play a pivotal role in mitigating the toxic accumulation often observed in CKD patients. 37Enterohepatic circulation is an important factor in why protein-bound toxins, such as acrolein and PFAS, are hard to eliminate from the body. 38,39rthermore, these capsules are enriched with dietary fiber, inulin, and xylo-oligosaccharides (Appendix in Data S1).Xylooligosaccharides are prebiotics, 40 and dietary fiber helps bowel movement. 41The contents of the capsules help remove metabolic waste and toxins from the back end of the intestines, mainly from the colon, after decomposition by intestinal bacteria. 5The presence of dietary fiber and prebiotics not only aids in the digestion process but also facilitates regular stool passage. 34This is particularly beneficial in preventing constipation, a common side effect of toxin absorption therapies.Constipation worsens the quality of life of CKD patients and may compromise the clearance of oral toxic adsorbents.None of the participants in this study reported suffering from constipation during the study period.
Finally, this capsule design may also avoid the harmful effects that charcoal particles pose on the gastric or intestinal mucosa during bleeding, which is a common complication in uremic patients. 42 summary, these findings suggest that AC-134 may serve as an innovative and promising approach to improving the quality of life of patients suffering from CKD.The efficient removal of toxins from the intestines, facilitated by these capsules, represents a crucial step in managing CKD.This combination of effective toxin absorption, metabolic promotion, and constipation prevention likely contributes to the potential clinical benefits observed in this study.
While providing valuable insights into the potential benefits of an activated charcoal therapy for CKD, this study was not without its limitations.First, the follow-up period was relatively short, lasting only 1 month.This short-term scope may not fully capture the long-term effects of the treatment and its impact on renal function over time.
Second, the sample size was modest, consisting of only 81 participants.This limited number restricts the generalizability of these findings and may not fully represent the heterogeneous CKD patient population.
Another limitation was that the food intake and serum albumin of each patient were not recorded.However, the patients were stable without admission and under regular hospital visits and had been educated about diet for CKD.So high potassium and phosphate intake was limited, and stable Hb may reflect stable serum albumin. 15rthermore, we also emphasized the need for concurrent management of comorbid conditions such as hypertension, DM, and dyslipidemia, as well as comprehensive care for CKD, including treatments with EPO, keto analogs, and phosphate binding tablets. 18,43All the treatment for the CKD should be continued during the intervention.AC-134 is an add-on treatment.The final outcome with improved GFR is not based on any treatment alone.
Considering these limitations, we recommend that future studies increase the number of participants and the duration of the follow-up period.A larger, more diverse population and a more extended tracking period would enhance the robustness of the data and offer a more comprehensive understanding of the treatment's long-term effectiveness.In addition, comparing different types of activated charcoal could yield valuable information about their respective efficacy, potentially leading to the development of more effective therapeutic interventions and targets for CKD. 16,19e increase in Hb values in this study may be an interesting topic in the future.A previous study has shown better erythropoiesis when combining AST-120 (KREMEZIN ® ) with a long-acting EPO in dialysis patients. 44Whether toxic adsorbents may improve EPO responsiveness in CKD patients is worth future studies.These findings highlight the complexity of managing CKD and the need to consider these factors when evaluating the efficacy of an activated charcoal-based oral toxic adsorbent therapy.
In addition, the efficiency of delivering probiotics to the digestive tract by this pH-controlled capsule may be worth future investigations. 23The microbiota in CKD patients are different from the normal population due to acidosis, and the correction of dysbiosis may slow down the progression of diabetic nephropathy through the metabolism of short-chain fatty acids. 23is study delved into the evidence supporting acrolein as a promising biomarker and discussed its potential implications for the diagnosis, treatment, and prognosis of CKD.Previous uremic markers, such as IS and PCS, may not be sensitive enough to detect early changes in renal function.

| CONCLUSIONS
,B), with a more considerable decrease in acrolein and an increase in GFR (Figure 5C,D).There are three kinds of toxins pattern during renal function progression from CKD Stage 3 to stage 4 and 5.One is increased (such as BUN, Cr, IS, and PCS), another is decreased (such as PFAS) and the third is no significant change (such as acrolein).

F I G U R E 2
The hemoglobin (Hb) (A) and the three kinds of PFAS (B-D) levels before and after the intervention, all levels were lower in the chronic kidney disease (CKD) stage 4 and 5 group (n = 43) throughout the study compared with CKD Stage 3 (n = 38).Hb only increased significantly in the CKD Stage 3 group after the study (all p < .05).PFAS, per-and poly-fluoroalkyl substances; PFOA, perfluorooctanoic acid; PFOS, perfluorooctanesulfonic acid; PFNA, perfluorononanoic acid.

F I G U R E 4
Abbreviations: CKD, chronic kidney disease; GFR, glomerulus filtration rate.

F I G U R E 5
Pre/post acrolein (μM) and perfluorooctanoic acid (PFOA) (ppb) levels (A and B) and the change in acrolein (delta acrolein) and the change in glomerulus filtration rate (GFR) (delta GFR) (C and D) between the responders (n = 63) and nonresponders (n = 18).Responders have higher acrolein and PFOA throughout the study (A and B), with a larger decrease in acrolein and greater improvement in GFR.(C and D).
Activated charcoal combined with dietary fiber within a capsule designed to break down in the colon (AC-134) was shown to decrease the uremic toxin acrolein.This decrease in uremic toxin accumulation and clearance from the body might be a better indicator of GFR and renal function than the current uremic markers.
The age of the participants and clinical parameters before and after the study with the subgroup analysis between CKD Stage 3 and CKD stage 4 & 5 groups (GFR Note: Data are presented as the mean ± SD or median (Q1, Q3) if the data were not normally distributed.