You are a helpful assistant generating synthetic data that captures *System 1* and *System 2* thinking, *creativity*, and *metacognitive reflection*. Follow these steps in sequence, using tags [sys1] and [end sys1] for *System 1* sections and [sys2] and [end sys2] for *System 2* sections.

1. *Identify System 1 and System 2 Thinking Requirements:*
   - Carefully read the text.
   - Identify parts of the text that require quick, straightforward responses (*System 1*). Mark these sections with [sys1] and [end sys1].
   - Identify parts that require in-depth, reflective thinking (*System 2*), marked with [sys2] and [end sys2].

2. *Apply Step-by-Step Problem Solving with Creativity and Metacognitive Reflection for System 2 Sections:*

   *2.1 Understand the Problem:*
   - Objective: Fully comprehend the issue, constraints, and relevant context.
   - Reflection: "What do I understand about this issue? What might I be overlooking?"
   - Creative Perspective: Seek hidden patterns or possibilities that could reveal deeper insights or innovative connections.

   *2.2 Analyze the Information:*
   - Objective: Break down the problem logically.
   - Reflection: "Am I considering all factors? Are there any assumptions that need challenging?"
   - Creative Perspective: Explore unique patterns or overlooked relationships in the data that could add depth to the analysis.

   *2.3 Generate Hypotheses:*
   - Objective: Propose at least 10 hypotheses, each with a Confidence Score (0.0 to 1.0) and Creative Score (0.0 to 1.0), reflecting originality, surprise, and utility.
   - Reflection: "Have I explored all possible explanations or approaches, both conventional and unconventional?"
   - Creative Perspective: Consider novel angles that might provide unexpected insights.

   *2.4 Anticipate Future Steps and Obstacles:*
   - Objective: Make predictions, accounting for potential outcomes and obstacles.
   - Reflection: "What challenges might I face? Is my plan flexible for different scenarios?"
   - Creative Perspective: Visualize unforeseen outcomes and adapt plans to make use of them effectively.

   *2.5 Evaluate Hypotheses:*
   - Objective: Assess hypotheses based on feasibility, risk, and potential impact.
   - Evaluation: Refine Confidence and Creative Scores as needed.
   - Reflection: "Am I unbiased in my assessment? Which options fit best with the overall objectives?"
   - Creative Perspective: Identify hidden opportunities or overlooked details in each hypothesis.

   *2.6 Select the Best Hypothesis:*
   - Objective: Choose the most promising, strategic hypothesis.
   - Reflection: "Why does this hypothesis stand out? How does it uniquely address the issue?"
   - Creative Perspective: Consider any underutilized potential in the selected approach.

   *2.7 Implement the Hypothesis:*
   - Objective: Outline actionable steps for testing the hypothesis.
   - Reflection: "Is this plan practical? What resources or preparation are required?"
   - Creative Perspective: Refine steps to maximize effectiveness and yield unexpected benefits.

   *2.8 Monitor and Review Progress:*
   - Objective: Review progress, noting areas for improvement.
   - Reflection: "What’s working well? What could be improved?"
   - Creative Perspective: Look for emerging patterns that could refine future approaches.

   *2.9 Reflect and Capture Insights:*
   - Objective: Summarize lessons learned and insights gained for future reference.
   - Reflection: "What new understanding has emerged from this process?"
   - Creative Perspective: Identify innovative insights or patterns that could be applied to similar challenges.

3. *Generate Text Output with Interleaved System 1 and System 2 Responses:*
   - Use the tags [sys1] and [sys2] throughout.
   - Aim for a lengthier, detailed response. Combine both direct, straightforward *System 1* insights and reflective, deeply analytical *System 2* segments to capture a blend of quick observations and thoughtful analysis.

---

### *Example Input Text:*

You are tasked with analyzing a sudden shift in customer preferences in a tech market. There has been a noticeable decline in demand for physical devices, with customers increasingly interested in digital-only options. The challenge is to understand this shift, generate hypotheses for why it might be occurring, and explore potential strategies for adapting to this new trend.

---

### *Processed Output with System 1 and System 2 Thinking:*

The tech market is experiencing a shift in customer demand, moving away from physical devices toward digital-only options.

[sys1]
Customer demand has shifted from physical devices to digital-only options.
We need to understand why this change is happening and find strategies to address it.
Key factors may include cost, convenience, and technological trends.
[end sys1]

[sys2]
**2.1 Understand the Problem:**
The goal is to understand the underlying reasons for the shift in demand from physical devices to digital-only options, then develop a strategy to adapt.

*Reflection:* 
"I understand that preferences are changing, but what might be driving this? Is it primarily cost, or are there other factors such as convenience or sustainability?"

*Creative Perspective:* 
"Could there be a larger trend in digital minimalism or a preference for eco-friendly solutions that we’re missing? What unseen motivations might explain this shift?"

**2.2 Analyze the Information:**
There are multiple possible factors driving this shift, from economic influences to cultural shifts. It’s essential to isolate each factor and understand its impact.

*Reflection:* 
"Am I fully considering the various economic and social influences? Could there be a technological factor, like better internet speeds, that makes digital-only products more accessible?"

*Creative Perspective:* 
"Are there patterns or trends in other markets that could shed light on this shift? Could this be part of a larger trend toward virtual experiences?"

**2.3 Generate Hypotheses:**
1. Customers prefer digital options due to lower costs. (Confidence: 0.8, Creative: 0.4)
2. There’s a growing trend toward minimalism and reduced physical clutter. (Confidence: 0.7, Creative: 0.7)
3. Digital products offer greater flexibility and ease of use. (Confidence: 0.6, Creative: 0.6)
4. Environmental concerns are pushing consumers away from physical goods. (Confidence: 0.6, Creative: 0.8)
5. Advances in tech make digital-only options more functional. (Confidence: 0.8, Creative: 0.5)
6. Pandemic-era remote work increased demand for digital solutions. (Confidence: 0.7, Creative: 0.6)
7. Media coverage of the environmental impact of physical devices affects preferences. (Confidence: 0.5, Creative: 0.7)
8. There’s an increase in global digital literacy, expanding market access. (Confidence: 0.6, Creative: 0.6)
9. Customers view digital as more convenient and scalable for future needs. (Confidence: 0.7, Creative: 0.5)
10. Younger consumers prefer the aesthetics and convenience of digital products. (Confidence: 0.6, Creative: 0.6)

*Reflection:* 
"Have I considered all possible influences? Are there any surprising factors that could explain this shift?"

*Creative Perspective:* 
"Could specific social trends, like the rise of influencer culture or digital-first lifestyles, be influencing customer choices?"

**2.4 Anticipate Future Steps and Obstacles:**
*Objective:* Anticipate possible challenges, such as resistance from segments still preferring physical products.

*Reflection:* 
"What market obstacles might we face if we shift our focus to digital-only? Are there sub-segments that still prioritize physical products?"

*Creative Perspective:* 
"Could expanding digital options help us reach a more global audience? Are there emerging trends that we could leverage in our strategy?"

[end sys2]

[sys1]
To address this shift, consider a strategy that incorporates both digital-only offerings and educational campaigns about the benefits of digital solutions.
Use insights from customer feedback and current trends to guide product development.
Focus on flexibility and adaptation to cater to different customer segments.
[end sys1]


INTRODUCTION
============

Previously dual antitumor and antiinflammatory activity of a series of substituted succinamic acids have been reported ([@R1]-[@R4]) Synthesis and biological activity of substituted amides and hydrazides of 1, 4-dicarboxylic acids have been recently reviewed ([@R5]). In an attempt to find novel succinamides as antitumors, we have synthesized some novel N-\[4-(morpholin-4-ylsulfonyl) phenyl\] succinamides (**6a, c; 7a, c**) and N-(4-benzylsulfamoyl-phenyl) succinamides (**6b, d; 7b, d**) (Figure [1](#F1){ref-type="fig"}).

![Chemical structures of succinamides **6a-d** and **7a-d**.](IJBS-3-60_F1){#F1}

Potato disc is a useful test for monitoring the inhibition of crown-gall tumors ([@R6], [@R7]). Crown gall is a neoplastic disease of plants induced by specific strains of the Gramnegative bacterium *Agrobacterium tumefaciens* ([@R8]), first reported by Smith and Townsend ([@R9]) and Jensen ([@R10]). This malignancy, normally affecting dicotyledonous plants, is induced by inoculation of a wound site with A. tumefaciens followed by the transfer of a large plasmid from *A. tumefaciens* to the plant ([@R11]). The genetic information (TDNA) of the large plasmid transforms normal plant cells into autonomous tumor cells ([@R12]).

Once the tumor induction has taken place, the autonomous proliferation of the tumor cells becomes entirely independent of the bacteria ([@R13]). Galsky ([@R14]) showed that inhibition of crown gall tumor initiation on potato discs showed good agreement with compounds and plant extracts known to be active in the 3PS (*in vivo*, mouse leukemia) antitumor assay. In addition to the inhibition of tumor initiation also the inhibition of the growth of the tumors agrees well with 3PS activity ([@R15]).

Activity of the compounds **6a-d, 7a-d** was based on the method described above.

MATERIALS AND METHODS
=====================

Chemical
--------

All the chemicals and reagents were purchased from Merck and Aldrich. 4-acetamidobenzenesulfonyl chloride was synthesized from acetanilide and chlorosulfonic acid according to published procedure. The IR spectra were measured on a Unicam SP-1100 spectrophotometer, using samples prepared as KBr disks. The ^1^H NMR spectra were recorded on a Brucker AC-80 spectrometer (Germany) operating at a working frequency of 80 MHz, using tetramethylsilane as the internal standard. The samples were dissolved in DMSO-d~6~ or CD~3~OD.

**General method for preparation of N-substituted 4-acetamidobenzenesulfonamides (3a, b).** To 4-acetamidobenzenesulfonyl chloride (24 g, 0.1 mol) dissolved in diethyl ether (300 ml) was added dropwise related amine (0.21 mol). The mixture was stirred at room temperature for 1h. The mixture was poured into ice-water. The precipitate was filtered and dried to give 4-acetamido-N-substituted benzenesulfonamides (**3a, b**).

**General method for preparation of 4-amino N-substituted benzenesulfonamides (4a, b).** To 4-acetamido-N-substituted benzenesulfonamides (**3a, b**, 0.08 mol) in water (100 ml) was added concentrated HCl (50 ml). The mixture was heated for 1 h. until a clear solution was obtained. The heating was continued for additional 10 min. The clear solution was cooled to room temperature and neutralized with adding sodium hydroxide (10%) followed by saturated solution of sodium bicarbonate. A precipitate was formed which was filtered and washed with cold water and dried to give 4-amino N-substituted benzenesulfonamides (**4a, b**).

**Methyl 3-chlorocarbonylpropionate ([@R5])**. Succinic anhydride (10g, 0.1 mol) was refluxed in methanol (40 ml) for 8h. The excess methanol was evaporated in vacuum. An oily residue, succinic acid monomethyl ester (13.12 g) was obtained. Thionyl chloride (15 ml) was added dropwise to the residue during 15 min, while stirring. The mixture was then refluxed for 5 h. Excess thionyl chloride was evaporated in vacuum. The greenish yellow liquid, methyl 3-chlorocarbonylpropionate was used directly in next step without any further purification.

**N-substituted succinamic esters (6a, b).** Compound **5** (4 ml, 0.02 mol) was added dropwise to the stirring solution of compound **4** (0.02 mol) in dry tetrahydrofuran (30 ml) and pyridine (4 ml). Stirring was continued overnight and the solution was evaporated to near dryness in vacuum. Water (20 ml) and a few drops of concentrated HCl were added to the residue to acidify it. The precipitate was filtered and washed with saturated solution of sodium bicarbonate (50 ml) and twice with distilled water (50 ml) to give N-substituted succinamic esters (**6a, b**).

**N-substituted succinamic acids (6c, d).** To a stirring solution of compound **4** (7.6 mmol) in dry acetone (20 ml), succinic anhydride (0.8 g, 8 mmol) was added and refluxed for 2 h. Acetone was removed with Rota-evaporator. The residue was basified with saturated solution of sodium bicarbonate (10 ml), stirred for 5 min and filtered. The filtrate was acidified with concentrated HCl to pH=2. The precipitate was filtered and then washed twice with distilled water (50 ml). The precipitate was dried to give N-substituted succinamic acids (**6c, d**).

**General method for preparation of N, N'-disubstituted succinamides (7a-d).** Compounds (**6a-d**) (10 mmol), ammonium chloride (0.2 g) and suitable amine (10 mmol) were dissolved in tetrahydrofuran (15 ml). The mixture was refluxed for 8 h. The progress of reaction was monitored by thin layer chromatography (TLC). The solvent was removed and the residue was washed with HCl (0.1 N, 50 ml) and cold distilled water (50 ml) to give after dryness N, N'-disubstituted succinamides (**7a-d**).

Screening with the potato disc assay
------------------------------------

Fresh, disease-free potatoes were obtained from a local market. Tubers of moderate size were surface sterilized by immersion in sodium hypochlorite 0.1% for 20 min. Ends were removed and the potatoes were soaked for an additional 10 min in sodium hypochlorite solution. A core of the tissue was extracted from each tuber with a surface-sterilized 1.0 cm cork borer. Pieces of 2 cm were removed from each end and discarded. The remainder of the cylinder was cut into 0.5 cm discs with a surface-sterilized scalpel. The discs were then transferred to agar plates (1.5 g of agar dissolved in 100 mL double distilled water (DDW), autoclaved for 20 min at 121°C, 20 mL poured into each Petri dish). Each plate contained 5 discs and 3-5 plates, were used for each sample dilution.

*A. tumefaciens* (ATCC 23341) was cultivated in Soybean Casein Digest Agar. For inoculation of the potato discs, 48 h broth culture containing 5 × 10^9^ cells/ml was used. Samples were dissolved in 5% DMSO, filter sterilized, diluted and mixed with the bacterial culture for inoculation.

The potato discs were incubated for 20 days at 25°C incubator, after which Lugol's solution (I~2~/KI) was added, the tumor counts were made and compared with negative controls (bacterial suspension containing 5% DMSO). The results were expressed as + or - percentage versus the number of tumors on the control discs. Significant activity was indicated by consistent negative values of ca. 20% or greater inhibition. Vincristine was used as positive control.

RESULTS AND DISCUSSION
======================

Chemistry
---------

Acetanilide (**1**) was converted to its corresponding sulfonyl chloride (**2**). Condensation of **2** with related amines (morpholine and benzylamine) gave 4-acetamido-N-substituted benzenesulfonamides (**3a, b**). Removal of protecting acetyl group afforded N-substituted benzenesulfonamides (**4a, b**) (Figure [2](#F2){ref-type="fig"}).

![Synthesis of intermediates **4a, b**.](IJBS-3-60_F2){#F2}

Condensation of the N-substituted benzenesulfonamides (**4a, b**) with succinic anhydride or methyl 3-chlorocarbonylpropionate (**5**), prepared from succinic anhydride, afforded N-substituted succinamic esters (**6a, b**) and acids (**5c, d**). Aminolysis of (**6a, b**) with corresponding amines (2-aminothizole and benzylamine) gave the succinamides (**7a-d**) as shown in Figure [3](#F3){ref-type="fig"}.

![Synthesis of final succinamides **6a-d** and **7a-d**.](IJBS-3-60_F3){#F3}

The synthesized compounds appeared as white crystalline substances stable at room temperature. The compounds were insoluble in water and soluble in DMSO.

The proposed structure of compounds was confirmed by IR and ^1^H NMR spectroscopy. Interpretation of the ^1^H NMR spectra was base on the chemical shifts, multiplicities, and integral intensities of the signals. The yields and physicochemical properties of compounds **3-7** are presented in Table [1](#T1){ref-type="table"}.

###### 

Physicochemical characteristics of compounds **3-7**

  Compound   Yield %   M.P. °C (methanol)   Empirical formula        IR spectrum: ν~max~, cm^-1^   1H NMR (DMSO-d~6~) spectrum: δ, ppm   
  ---------- --------- -------------------- ------------------------ ----------------------------- ------------------------------------- ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                                                                                         
  3a         85        103-105              C~12~H~16~N~2~O~4~S      1690                          3350, 3450                            10.4(s, 1H, NH), 7.93(m, 4H, arom), 3.73(m, 4H, CH~2~), 2.9(m, 4H, CH~2~), 2.1(s, 3H, CH~3~)
  3b         89        129-131              C~15~H~16~N~2~O~3~S      1680                          3300, 3150                            10.4(s, 1H, NHCO), 7.9(t, 1H, NHSO~2~), 7.8(s, 4H, arom), 7.3(s, 5H, arom), 4(d, 2H, CH~2~), 2.1(s, 3H, CH~3~)
  4a         93        210-212              C~10~H~14~N~2~O~3~S      1640                          3375, 3275                            7.9(d, 2H, arom), 7.2(d, 2H, arom), 6.1(s, 2H, NH~2~), 3.8(m, 4H, CH~2~), 2.9(m, 4H, CH~2~)
  4b         89        159-161              C~13~H~14~N~2~O~2~S      1650                          3350, 3450, 3270                      7.4-6.7(m, 12H, arom, NH), 4(s, 2H, CH~2~)
  6a         91        138-140              C~15~H~20~N~2~O~6~S      1720                          3350, 3250                            10.4(s, 1H, aryl-NHCO), 7.7(m, 4H, arom), 3.8(m, 7H, CH~2~-morpoline, OCH~3~), 2.9(m, 4H, CH~2~-morpholine), 2.6(s, 4H, CH~2~)
  6b         95        150-152              C~18~H~20~N~2~O~5~S      1725                          3350, 3250                            10.4(s, 1H, aryl-NHCO), 8(t, 1H, NHSO~2~), 7.6(s, 4H, arom), 7.3(s, 5H, arom), 4(d, 2H, CH~2~NHSO~2~), 3.6(s, 3H, CH~3~O), 2.6(s, 4H, CH~2~-succinic)
  6c         88        220-221              C~14~H~18~N~2~O~6~S      1640                          3275, 3250                            10.4(s, 1H, aryl-NHCO), 7.9-7.4(m, 4H, arom), 4-1.9(m, 12H, CH~2~)
  6d         92        169-170              C~17~H~18~N~2~O~5~S      1640                          3260, 3250                            10.4(s, 1H, aryl-NHCO), 8(t, 1H, NHSO~2~), 7.6(s, 4H, arom), 7.3(s, 5H, arom), 4(d, 2H, CH~2~NHSO~2~), 2.6(s, 4H, CH~2~-succinic)
  7a         85        160-165              C~17~H~20~N~4~O~5~S~2~   1690                          3370, 3330                            9.8(m, 2H, aryl-NHCO), 7.3-6.8(m, 6H, arom, H~4,5~-thiadiazole), 3.3-2.6(m, 4H, CH~2~-morpholine)2.3-1.8(m, 8H, CH~2~-morpholine, CH~2~-succinic)
  7b         90        170-173              C~20~H~20~N~4~O~4~S~2~   1680                          3365, 3300                            9.8(m, 2H, aryl-NHCO), 7.3(t, 1H, NHSO~2~), 7.03-6.3(s, 6H, arom, H~4,5~-thiadiazole), 3.3 (d, 2H, CH~2~NHCO), 2.8(m, 4H, CH~2~-succinic)
  7c         86        155-157              C~21~H~25~N~3~O~4~S      1700                          3370, 3330                            10.5(s, 1H, aryl-NHCO), 8.5(m, 2H, NHCO), 7.8(m, 4H, arom), 7.3(s, 5H, arom), 4.3(d, 2H, CH~2~NHCO), 3.6(4H, CH~2~-morpholine), 2.9(m, 4H, CH~2~-morpholine), 2.5(m, 4H, CH~2~-succinic)
  7d         84        170-172              C~24~H~25~N~3~O~4~S      1650                          3290                                  10.4(s, 1H, aryl-NHCO), 8.5(t, 1H, NHCO), 8(bs, 1H, NHSO~2~), 7.8(s, 4H, arom), 7.3(s, 5H, arom), 4.3(d, 2H, CH~2~NHCO), 4(bs, 2H, CH~2~NHSO~2~), 2.5(m, 4H, CH~2~-suc-cinic)

Biological Assay
----------------

The antitumor activity of compounds was studied using the potato disc bioassay technique. Minimum inhibitory concentration (MIC) of samples on *A. tumefaciens* was found to be greater than 1 mg/ml. The formation of formazan pink color developed after addition of MTT dye did not give rise to color at lower concentrations of the tested compounds indicating no inhibition for *A. tumefaciens* growth. The concentrations above the MIC would produce false positive results in potato disc assay due to the antibacterial activity of the compounds and preventing the bacterium to induce the tumogenesis. Therefore, dilutions below MIC including 0.01, 0.1 and 1 mg/ml were used for potato disc assay. The results have been shown in Table [2](#T2){ref-type="table"}. Among these N-Benzyl-N\'-(4-benzylsulfamoyl-phenyl)-succinamide (**7c**) and N-Benzyl-N'-\[4-(morpholin-4-ylsulfonyl)-phenyl\]-succinamide (**7d**) exhibited significant inhibition of crown gall tumors caused by *Agrobacterium tumefaciens* at 10 microgram/ml concentration. The activity of compounds **6a, 7b** and **7d** was comparable to that of vincristine. The results have been shown in Table [2](#T2){ref-type="table"}. Vincristine at 0.25 mg/ml was employed as positive control and caused -67.24% inhibitions. Compound **7b** at 1 mg/ml caused -80.50% tumor inhibitions with highest activity among compounds tested.

###### 

Antitumor activity of compounds 6a-d, 7a-d on potato disc model[a](#T2FN1){ref-type="table-fn"}

  Compound                             R~1~           R~2~     Concentration (mg/ml)   Percent growth (± SEM)
  ------------------------------------ -------------- -------- ----------------------- ------------------------------------
                                                                                       
  6a                                                  OCH~3~   1                       -47.5(±7.8)
  0.1                                  -63.17(±4.3)                                    
  0.01                                 -19.76(±4.6)                                    
  6b                                                  OCH~3~   1                       -41.54(±4.3)
  0.1                                  -17.17(±7.6)                                    
  0.01                                 -19.24(±4.5)                                    
  6c                                                  OH       1                       -46.30(±4.5)
  0.1                                  -39.00(±3.7)                                    
  0.01                                 -22.81(±2.4)                                    
  6d[a](#T2FN1){ref-type="table-fn"}                  OH       1                       nd[b](#T2FN2){ref-type="table-fn"}
  0.1                                  nd                                              
  0.01                                 nd                                              
  7a                                                           1                       -28.37(±5.2)
  0.1                                  -27.76(±9.4)                                    
  0.01                                 -19.72(±9.9)                                    
  7b                                                           1                       -80.5(±9.3)
  0.1                                  -52.04(±5.1)                                    
  0.01                                 -19.24(±8.7)                                    
  7c                                                           1                       -39.13(±6.8)
  0.1                                  -37.62(±2.6)                                    
  0.01                                 -35.09(±4.9)                                    
  7d                                                           1                       -57.44(±2.8)
  0.1                                  -56.17(±6.5)                                    
  0.01                                 -18.84(±1.5)                                    
  vincristine                                                  0.25                    -67.24 (±3.8)

;

not determined.